quantitative studies of auditory hair cells and nerves in lizards

h

Quantitative Studies of Auditory Hair Cells and Nerves in Lizards

MALCOLM R. MILLER Department of Anatomy, University of California, San Francisco

San Francisco, California 94143

ABSTRACT Because the lizard cochlear duct is anatomically accessible as well as

relatively simple in structure it is an excellent model in which to study auditory hair cells, nerve fibers, and innervational patterns. The objectives of this study were to determine the intra- and interspecific variations of auditory hair cell and nerve fiber numbers, nerve fiberhair cell ratios, and nerve fiber sizes in a variety of lizard species and to relate these to auditory function and phylogeny.

Hair cell numbers were determined by SEM and serial frontal sections of the papilla basilaris and nerve fiber numbers and diameters by use of a Zeiss TGZ3 particle counter. The coefficient of variation of hair cell numbers varied from 3.2 to 16.6 (171 specimens, 15 species) and of nerve fiber numbers from 1.2 to 14.4 (381 specimens, 35 species). There was no correlation between hair cell or nerve fiber number and age or sex.

The nerve fiber numberhair cell number ratio was 3.5-11.1/1 in small papillae basilares of the iguanid-agamid-anguid type, 2.4-3.211 in the teiid type, and 0.6-1.5/1 in the larger specialized papillae of the scincid and gekkonid types.

Nerve fibers varied in diameter from 0.8 to 6.0 pm (largest percentage were 2-4 pm) and were unimodally distributed. Larger nerve fibers usually supplied the unidirectionally oriented hair cells of the papilla basilaris.

Variations in hair cell and nerve fiber numbers in other vertebrate classes and the functional and phylogenetic aspects of lizard papilla basilaris structure and innervation are discussed.

Key words: papilla basilaris, auditory nerve, reptile ear, auditory hair cellheme fiber ratios, nerve fiber diameters

The numbers of auditory hair cells or nerve fibers have been reported in a variety of vertebrate species (fish, Cor- win, '81, '83; Fay and Popper, '80; amphibians, Lewis et al., '84; Dunn, '78; reptiles, Wever, '78; Weiss et al., '76; Bagger- Sjoback, '76; Leake, '77; birds, Boord, '69; mammals, Ras- mussen, '40; Boord and Rasmussen, '58; Bredberg, '68; Ga- cek and Rasmussen, '61; Ehret and Frankenreiter, '77; Ehret, '79; Morrison et al., '75; Ramprashad et al., '78; Retzius, 1884; Wever et al., '71, '72). With few exceptions (Rasmussen, '40; Bredberg, '68; and Dunn, '781, these reports are based on relatively few specimens and there are no studies of both hair cell and nerve fiber numbers in the same animal. Since auditory function is probably closely related both to the number of sensory units and their innervating fibers, it is important to know the normal ranges in hair cell and nerve fiber numbers as well as the proportions of nerve fibers to hair cells and particularly the precise mode of hair cell innervation.

The lack of more complete studies of hair cell and nerve fiber numbers is due to the anatomical inaccessibility of the cochlea and to the relatively large numbers of sensory cells and innervating fibers, especially in mammals.

In birds and the crocodilian and testudinate reptiles, the cochlear duct is much more accessible than in mammals, but their large numbers of hair cells and nerve fibers also make quantitation difficult. Saurian reptiles (lizards and snakes), particularly lizards with small papillae basilares, are most amenable to quantitative study. In amphibians both the papilla basilaris and the papilla amphibiorum contain sufficiently few hair cells to be counted accurately, but Lewis and Li ('73) suggest that hair cells are added to these papillae during growth of the animal, and thus hair

Accepted August 28, 1984.

0 1985 ALAN R. LISS, INC.

2 M.R.MILLER

cell number is dependent on age and animal size. Similarly in fishes it has been shown that auditory hair cells increase markedly with aging (Corwin, '83; Fay and Popper, '80).

Compared to that of other vertebrates, not only is the cochlear duct in lizards easily exposed and fixed, but also many species have relatively small numbers of hair cells and innervating fibers so that precise quantitation is possible. Furthermore there is 110 evidence for growth of the auditory papilla after birth (Miller, '66), and lizard cochleae exhibit a variety of structurally and functionally different types (Miller, '80; Manley, '81).

MATERIALS AND METHODS Previous studies of the auditory papillae of lizards (Miller,

'80, '81) provided a base from which to select species that ranged from those considered simpler to more complex types. Species from ten families with different types of papillar structure were chosen for study, although selection was often determined by availability. Recent changes in state, federal, and foreign legislation have made it very difficult or impossible to obtain some reptilian species.

Iguanid lizards comprise a large part of the total sample, because several sub-groups in this family show significant structural differences in the auditory papilla (Miller, '81). Also, many iguanine and sceloporine iguanids were studied because the smaller numbers of papillar hair cells in these species make them ideal for quantitative investigation.

Reliable quantitative data require much larger sample sizes than have been reported in the literature on the sub- ject of variation in hair cell and nerve fiber numbers in auditory organs. Therefore as many specimens as possible were examined so that sufficient data for statistical evaluation would be available. Altogether, 37 species of lizards representing ten families were studied (Table la,b).

For study of either the papilla basilaris or the papillar nerve, animals were decapitated, the otic capsules were opened as rapidly as possible (Miller, '81), and the entire head was immersed in a variety of fixatives. When the papilla was to be examined by scanning microscopy, heads were fixed in 3% glutaraldeliyde in 0.1 M cacodylate buffer. Subsequent processing for SEM study is described by Miller ('81). For histological study of the papilla basilaris or the papillar nerve by serial thick sections (1 pm thickness), a number of fixatives using various combinations of acrolein, glutaraldehyde, and paraformaldehyde were used. After many trials it was found that for studying both papilla and nerve the best fixative was 0.5% acrolein and 2.5% glutaraldehyde in 0.1 M cacodylate buffer. Cochlear ducts were fixed intact in the opened otic capsule for 1-3 days at 4"C, and then dissected out of the capsule and placed in 1% EDTA in 2% glutaraldehyde for 3-4 days to decalcify the lagenar otolithic crystals. These specimens were then embedded in Epon-Araldite and serially sectioned in sagittal, transverse, or frontal planes.

The number of hair cells contained in the papilla basilaris was best determined by SEM or serial frontal sections of the papilla. With either method smaller papillae composed of 200 hair cells or less provided the most accurate data. When papillae with more than 200 hair cells were counted by SEM, the number of specimens with all hair cells dis- crete and unobscured (less than 5%) were too few to warrant the expense of obtaining sufficiently large samples by this method. Certain types of auditory papillae with hair cell numbers ranging from 200 to 500 (e.g., tropidurine iguanids, agamids, and diploglossine anguids) could be serially

sectioned in the frontal plane and accurate hair cell counts made. In papillae with more than 500 hair cells only a very few SEM specimens were sufficiently perfect to provide accurate hair cell counts. For good estimates of hair cell numbers in larger papillae a combination of SEM and serial sagittal sections were useful. The number of specimens of each species used to determine hair cell number is given in Table la .

Determination of papillar nerve fiber number is much easier and more certain than hair cell number. Hence the number of nerve fiber counts (Table lb) is considerably larger than the number of hair cell counts. In most specimens papillar nerves were serially sectioned at 1-pm intervals from the posterior ganglion to the point of entry of the nerve into the papilla basilaris. Nerve fiber numbers and diameters were determined by photographing an entire nerve cross section under oil immersion and subsequent prints were enlarged to x 1,550. Correct magnification was assured by concomitantly photographing a slide of a ruled scale. Each papillar nerve cross section was reconstructed as a montage consisting of three to 30 photographs depending on the size of the nerve. All nerve fibers were counted either by using a plastic (Saran) overlay and marking and tabulating each fiber, or by simultaneously counting and measuring the diameter of each nerve fiber with a Zeiss TGZ3 particle counter. In this way a total of 629,832 nerve fibers were counted in 652 complete montages from 414 specimens in 37 species of lizards (ten families).

Studies of serial sections of the entire papillar nerve showed that the most optimal site for nerve fiber measurement was 20 pm distal to the ganglion where the greatest number of nerve fibers were cut in the transverse (rather than the oblique) plane.

In 393 specimens the diameter of each papillar nerve fiber (including the myelin sheath) was measured with the particle counter. Where fibers were relatively circular in cross section, determination of maximum diameter posed no problem. In obliquely cut fibers, however, the largest diameter of the short axis was measured. Since the particle counter intervals of measurement were approximately 0.125 pm, these small increments were combined into nerve fiber diameters of 1-pm intervals. The numbers of specimens used to measure nerve fiber sizes and all data from these studies are listed in Table 3. Dr. David Heilbron (chief of the Computer Sciences Center at the University of Califor- nia, San Francisco) advised that it was legitimate to treat each 1-pm increment of nerve fiber measurements statisti- cally, and to combine the nerve fibers into two size groups (0.8-3 pm and 3-6 pm) since all but a very small percent of fibers occur within this range. These subdivisions were subsequently useful in relating nerve fiber sizes to different structural types of auditory papillae.

I am aware of'the potential errors inherent in measuring nerve fiber diameters due to such variable factors as fixa- tion, optics, photography, and idiosyncrasies of the observers. In these studies two different observers shared the task of measuring all nerve fibers in a total of 393 nerves which included measurements made at both single and multiple levels of the papillar nerve and a series of repeat counts to assess accuracy. A total of 379,638 nerve fiber diameters were measured.

Variations in measurements of nerve fiber diameters made either by the same observer or by two different observers were checked in the following way: all the nerve fibers in four identical copies of cross sections of nerves

LIZARD AUDITORY HAIR CELLS AND NERVES 3

TABLE 1. Variations in Pauillar Hair Cell Number and Nerve Fiber Number

C.V.' PNFNi

Species specimens Range Mean deviation ("," 1 specimens Range Mean deviation C.V.' HCN2 Standard No. of Standard --x No. of

Iguanidae (Anolines)

Anolis carolinensis Anolis equestris Polychrus marmoratus

(Iguanines) Crotaphytus wislizeni Crotaphytus collaris

Crotaphytus collaris

Dipsosaurus dorsalis Phrynosoma platyrhinus Sauromalus obesus

(Sceloporines) Callisaurus draconoides Uta stansburiana Sceloporus magister

(Tropidurines) Tropidurus hispidis Plica plica (aj3 Plica plica (6) Plica umbra Uranoscodon supraciliaris Leiocephalus schreibersi

Agama agama Acanthosaura crucigera

Celestus costatus Ophisaurus ventralis

hcer ta sicula (aj3 Lacerta sicula (b)

Ameiua ameiua Cnemidophorus motaguae Cnemidophorus tigris Callopistes maculatus

Varanus exanthematicus

Coleonyx uariegatus Cosymbotus platyurus Gekko gecko Hemidactylus fienatus

Eumeces laticeps Eumeces obsoletus Mabuya multifasciata Mabuya parrotetii

baileyi

collaris

Agamidae

Anguidae

Lacertidae

Teiidae

Varanidae

Gekkonidae

Scincidae

Cordvlidae

-

3 2 2

24 18

7

27 6

15

6 14 13

1 2 4 4 4 4

4 4

7 1

1 -

1 2 3 -

1

1 1 6 2

1 4 1 1

Cordylis uittifer 1 Xantusiidae

a. Haircell No.

181-186 257-265 137-140

48-72 97-132

103-125

81-106 54-70 120-149

70-76 50-58 78-98

219 351-390 295-370 304-358 230-280 85-124

281-310 280-3004

380-438 225

102 -

730 400-4454 450-476 -

l,80O4

435 8004

183.0 261.0 138.5

60.1 111.8

115.4

90.7 60.3

137.5

73.0 54.1 89.8

- 370.5 335.0 333.5 259.0 108.5

294.8 290.0

395.9 -

- -

-

422.5 464.7 -

-

- -

2,000-2,200 2,100

9404 - 9004 900 1,2504 - 1,200~ -

5955 -

700-8584 779

2514 -

- - -

5.1 10.7

7.3

6.2 5.8 8.5

2.3 2.5 5.4

-

40.6 28.6 24.2 18.0

14.0 11.5

20.8 -

- -

- - - -

-

- - - -

- ~

~

-

-

-

b. Nerve-fiber No.

- - -

8.5 9.6

6.3

6.8 9.6 6.2

3.2 4.6 6.0

12.1 8.6 9.3

16.6

4.7 4.0

5.3 -

- -

- - - ~

-

- - - -

- - - -

-

-

23 4 7

37 17

8

31 5

16

5 10 17

12 4 4 6 6 5

8 6

11 6

4 4

7 10 24 2

3

8 5

11 7

14 10 6 6

2

775-1,006 882.4 965-1,134 1,067.5 420-534 478.3

514-795 641.7 1,117-1,404 1,240.0

854-1,092 990.4

401-597 497.1 265-384 309.4 441-641 528.1

520-626 569.6 374-460 433.8 5 4 6 - 6 9 0 607.9

1,246-1,516 1,407.3 1,536-1,632 1,584.5 1,521-1,693 1,598.0 1,522-1.182 1,660.8 991-1,143 1,090.2 787-811 798.0

1,000-1,159 1,084.4 1,050-1,286 1,145.2

1,240-1,634 1,479.5 693-884 798.4

479-545 512.8 535-601 565.8

1,654-1,854 1,739.4 1,242-1,507 1,369.1 943-1,258 1,104.3 1,474-1,476 1,475.0

1,821-2.073 1,987.7

581-760 673.6 622-699 662.2 1,667-2,102 1,905.2 462-635 535.6

554-702 631.0 497-621 561.3 1,098-1,306 1,176.3 928-1,082 1,007.3

460-549 504.5

65.4 73.8 45.2

71.2 75.5

78.8

48.5 44.6 67.8

44.0 29.5 47.6

90.0 41.3 81.9 98.2 65.7

9.5

49.8 97.9

150.2 77.6

27.1 32.7

82.7 80.5 83.3 -

-

67.4 34.8

135.8 63.4

44.4 41.8 91.9 59.1

-

7.4 6.9 9.5

11.1 6.1

8.0

9.8 14.4 12.8

7.7 6.8 7.8

6.4 2.6 5.1 5.9 6.0 1.2

4.6 8.5

10.2 9.7

5.3 5.8

4.8 5.9 7.5

10.0 5.3 7.1

11.8

7.0 7.4 7.8 5.9

7.5

4.8 4.1 3.5

10.7 11.1

8.6

5.5 5.1 3.8

7.8 8.0 6.8

6.4 4.3 4.8 5.0 4.2 7.4

3.7 3.9

3.7 3.5

5.0 -

2.4 3.2 2.4

~

1.1

1.5 0.8 0.9 0.7

0.7 0.6 0.9 0.8

0.8

Xantusia uigilis 3 -_ - 12 352-443 381.5 28.5 . 1.1

'Coefficient of variation. 'Ratio of mean papillar nerve fiber number to mean hair cell No. 3Apparently two different populations. 'Estimates form frontal papillary sections and scanning micrography Wever, '78).

4 M.R. MILLER

from five different specimens were measured by each observer. Measurements on two of the four copies were made at least 6 months apart. The means of the diameters of all the nerve fibers in each of five intervals of 1 pm (20 means for each observer) were compared and an evaluation of the ability of each individual to reproduce his or her own results, as well as the differences between the two observers, were determined. T tests of the means revealed no difference at the 96% confidence level either between the observations made by one individual or in those made by both observers in 60% of the trials. In the other 40% of the trials the percentages of difference were slightly above the values necessary to achieve the 96% confidence level. Since the results of these studies do not claim to differentiate between small differences among nerve fiber size groups, and only large size differences have been shown to be related to papillar structure variations, greater accuracy of method was not required.

A study was also made in ten species that compared (in each species) the difference in nerve fiber sizes when they were measured at four different levels of a single nerve with the mean fiber sizes at one level of four different nerves. The mean nerve fiber size determined by either of these methods revealed no differences by the T test at the 96% confidence level.

OBSERVATIONS Structure of the auditory papilla

The structure of the reptilian papilla basilaris has been extensively studied in recent years and comprehensive descriptions and reviews published by Baird ('741, Wever ('781, and Miller ('80). For reader orientation, Figures 1-3 depict some gross aspects of the lizard cochlear duct, papilla basilaris, and the papillar branch of the auditory nerve. Figure 4 diagrammatically illustrates the major types of lizard auditory papillae. Subsequent figures illustrate the range of papilla basilaris structure by both scanning microscopy and frontal sections together with cross sections of the corresponding papillar nerves.

Hair cell number and description of papillae basilares studied (200 specimens, 36 species, ten

families; Table la) Regardless of the structural type or complexity of an

auditory papilla, hair cell number is closely correlated with the length of the papilla. The coefficient of correlation for the 36 species used in this study was .967. Papilla basilaris length, however, varied from one lizard family to another depending on the structural type and complexity of the papilla (Miller, '66). Within any one lizard family papilla basilaris length was related to the adult body size of the species in that family (Miller, '66). The coefficient of correlation (r) between species size and papilla length in some lizard families is as follows: in ten species of iguanids, r = 279, nine species of agamids, r = ,376, seven species of teiids, r = .797, 11 species of gekkonids, r = ,846, and ten species of scincids, r = .193 (taken from original data published by Miller ('66) and the r values calculated for this paper).

Since hair cell numbers are best determined in small auditory papillae (0.15-0.5 mm in length) species of lizards from the families Iguanidae, Agamidae, and Anguidae provided most of the data concerning variation in hair cell numbers in this study. Of the above three families, the iguanids are the best represented because they possess a greater variety of auditory papillae, and species of this family are readily available. In the family Iguanidae, stud-

ies of several anatomical features have delineated six different types (Zug, '71; Savage, '58; Etheridge, '64; Wever, '78). According to Zug ('71) these types are (1) anolines, (2) basiliscines, (3) iguanines, (4) sceloporines, ( 5 ) tropidurines, and (6) malagasians. Miller ('81) demonstrated that the auditory papillae of each of the first four above types could be distinguished from one another. The present study has reconfirmed the above findings, and in addition has made observations on several heretofore undescribed tropidurine iguanids.

In the following brief descriptions of the papillae basilares studied, sections of the corresponding papillar nerves are illustrated and where pertinent, the relationship of different sized groups of nerve fibers to specific regions of the papilla are noted.

Anoline iguanids (Table la, Figs. 5-7). The papilla basilaris of Anolis carolinensis has been previously described in detail (Miller, '811, but is illustrated in Figure 5 to demonstrate the apical patch of unidirectional hair cells and the elongated basal region made up of bidirectional hair cells. The frontal section of the papilla of Anolis equestris (Fig. 6) is very similar to that ofA. carolinensis but is larger and contains more hair cells. The longer papilla and larger hair cell number in A. equestris is related to the greater adult body size of this species. While Polychrus marmoratus is a larger species than Anolis carolinensis, its papilla contains fewer hair cells and the apical region consists of a larger proportion of undirectionally oriented hair cells (30% of the total) than does the apical region of A. carolinensis (4% of the total).

The papillae of Anolis carolinensis and A. equestris are more similar to those of the tropidurine iguanids, Tropidu- rus hispidis, Plicaplica, and Plica umbra than to the other species of iguanids so far studied (vide infra). The number of specimens of anoline iguanids in which hair cell number was determined is insufficient to establish hair cell number variability in this group.

A section of the papillar nerve of Anolis equestris (Fig. 7) shows that mostly larger nerve fibers supply the apically located unidirectional hair cells illustrated in the frontal section of the papilla (Fig. 6).

Zguanine and Sceloporine iguanids (Table la , Figs. 8- 10). The largest samples of animals studied are from these two iguanid sub-groups (130 specimens). The papillae of these types of iguanids are characterized by a centrally located group of tectorial-membrane-covered, short-ciliated, unidirectionally oriented hair cells and by apical and basal groups of long-ciliated, bidirectionally oriented hair cells with no tectorial covering (Fig. 8). Sceloporines have only two rows of bidirectional hair cells, while iguanines have three or more.

Section of the papillar nerve (Fig. 10) shows that larger nerve fibers supply the centrally located unidirectional hair cells (figs. 8,9).

A bbreuaations A apical (direction, anatomically antero-ventral) ar anterior root of auditory nerve g posterior ganglion of auditory nerve IV IVth cranial nerve (trochlear nerve) lg nerve to the macula lagenae mn nerve to the macula neglecta N neural direction (facing the incoming

innervating fibers) P papillar branch of the auditory nerve

(homologue of the cochlear nerve of mammals) pa nerve to the crista of the posterior ampulla pb pagilla basilaris V V' cranial nerve (trigeminal nerve)

LIZARD AUDITORY HAIR CELLS AND NERVES

Fig. 1. Stereophotographs of part of the brain and the attached cochlear ducts of the anguid lizard, Gerrhonotus multicarinatus. All cranial bones have been removed. The cochlear ducts are attached to the medulla by the posterior roots of the auditory nerve. x 10. From Miller ("75).

Fig. 2. Stereophotographs of the medial aspect of the left cochlear duct of Gerrhonotus multicart- natus showing the relationship of the posterior ganglion of the auditory nerve to the lagenar, papillar, and posterior ampullary nerve branches. x 18. From Miller ('75).

Fig. 3. Stereophotographs of the medial aspect of the left cochlear duct of the teiid lizard, Ameiua undulata (incorrectly labelled in Miller, '75) to illustrate one of the larger papillar nerves found in lizards. x 18. From Miller ('75).

5

6 M.R. MILLER

KlNOClLlAL ORIENTATION PAnERNS IN LIZARD PAPILLAE BASILAWS

NEURAL

f 4 (ventra1)APICAL + + EASAL(dorsa1)

ABNEURAL

IGUANID - AGAMlD - ANGUID'

Most iguonids Most ogamids Some agomids (Phrynocepholus. Uromastix. Physignaihus I (Gerrhonotus. Diploglossus. Anguids with reduced limbs Barislo) (Anguis. Ophisourus) %me iguonids (Anolids and

Pllca)

Anguids wiihoui limb reduction

LACERTID

TEND

VARANID A

GEKKONID

Fig. 4. Outline drawings of the papillar surface of several different types of lizard papillae basilares. The larger (heavier) arrows indicate unidirectionally oriented hair cells and the smaller arrows, bidirectionally oriented hair cells. The apex (or tip) of the arrows designates the side of the hair cell bearing the kinocilium which orients the cell physiologically. The papillae are not drawn to scale. From Miller ('80).

Thpidurine iguanids (Table la , Figs. 11-15). Tropidurus hispidis, Plica plica (Fig. ll), and Plica umbra (Fig. 12) have papillae similar to those found in the anoline iguanids in that the papillae are longer, hair cell numbers are larger (200400), and the unidirectional hair cells are located at the apical tip of the papilla. Although Uranoscodon supraciliaris has a longer papilla and more hair cells than the iguanines and sceloporines, like species of these sub-groups, it has centrally located unidirectional hair cells. The papilla basilaris of Leiocephalus schreibersi (Fig. 141, with a smaller number of hair cells (85-124) and centrally located unidirectional hair cells, is much more like the papillae of the iguanines and unlike those of Tropidurus and Plica. Figure 15 does not show a clear-cut segregation of larger nerve fibers supplying the unidirectional hair cell region of Leiocephalus. The larger number of hair cells in the first three species described above is related to the larger body size of these lizards and to the increase in the numbers of rows of bidirectional hair cells in their papillae.

The variations in papillar structure of the tropidurine iguanids reported here do not demonstrate the structural homogeneity so far observed in other iguanid sub-groups and indicate that the taxonomic relationships of Uranosce don and particularly of Leiocephalus need further investigation. From measurements of nerve fiber sizes, it was apparent that our specimens of Plica plica were from two different populations of animals, and they were thus sub-

divided into a and b groups. A similar situation was found in specimens of Lacerta sicula.

Agamidae (Table la). The papilla basilaris of Acanthe saura crucigera has not been described before but is very similar to that of Agama agama previously described in detail (Miller, '78b). Like most agamids (Fig. 4), the papillae of both these species are characterized by a small apical group of tectorial-membrane-covered unidirectional hair cells and a relatively long basal group of uncovered bidirectional hair cells. Sections of the papillar nerves of these species do not show a clear-cut distribution of larger-sized nerve fibers to the apical unidirectional hair cell region.

Anguidae (Table 1a; Figs. 16-20). Celestus costatus is a diploglossine type of anguid lizard with a papilla basilaris similar to that of the anoline and some types of tropidurine iguanids and to the agamids. The papilla of Celestus costatus (Figs. 16, 17) is one of the largest of the iguanid-agamid- anguid type (Fig. 4) and contains 380-438 hair cells. The tectorial-membrane-covered unidirectional hair cells are apical and the uncovered bidirectional hair cells basal in location. Gerrhonotus multicarinatus, an anguid lizard much studied by physiologists (Weiss et a]., '76; Holton and Hudspeth, '83), has a slightly smaller papilla (0.4 mm) than Celestus (0.5 mm) but contains only half the number of hair cells (Miller, '73b). The much larger number of hair cells in Celestus (Figs. 16,171 is accounted for by the presence of more rows (nine) of bidirectional hair cells than is found in Gerrhonotus (five). The papillar nerve of Celestus (Fig. 18) shows a very distinct distribution of larger nerve fibers to the apical unidirectional hair cell group.

On the other hand, Ophisaurus ventralis, which has been described before (Miller, '78b), is shown here (Fig. 19) to compare it with its nerve (Fig. 20) and with the papilla of Celestus. Like the iguanine and sceloporine iguanids (Fig. 41, Ophisaurus has centrally located unidirectional hair cells and fewer rows of bidirectional hail- cells than Celes- tus. Section of the papillar nerve of Ophisaurus ventralis (Fig. 20) demonstrates that some of the larger nerve fibers are related in position to the central unidirectional hair cell group of the papilla (Fig. 19).

Lucertidae (Table la). Unfortunately it was not possible to obtain a sufficient variety of species or numbers of these lizards to establish the variation in hair cell number. In general, lacertid species have small papillae with hair cell numbers ranging from 90 to 150 (Miller, '66, 78b, '80) and are characterized by papillae completely divided into two parts. A small group of unidirectionally oriented hair cells is located at the basal end of the basal segment and all other hair cells are bidirectionally oriented (Fig. 4). All hair cells are short-ciliated and are covered by a tectorial membrane. Nerve sections show that the larger nerve fibers enter the papilla at the location of the unidirectional hair cells.

In the species of the families described below, precise determination of hair cell number is very difficult because of greater papillar length and number of hair cells. What I consider to be accurate estimates of hair cell number in longer papillae (0.7-2.0 mm) were determined by a combination of SEM and serial sagittal sections of the papilla basilaris. It was necessary to have reliable estimates of hair cell numbers so that the nerve fiberhair cell ratio could be determined. As will be shown later, this ratio has significant functional and phylogenetic implications.

Teiidae (Table la, Figs. 21-23). The papillae basilares of Cnemidophorus tigris and Cnemidophorus motaguae (Figs. 21,221 are very similar to that of Ameiva ameiva which has been previously described (Miller, '73b). These papillae are

Fig. 5. SEM of the surface of the left papilla basilaris of Anolis carolinensis. The short-ciliated hair cells located apically (reader’s left) are unidirectionally oriented and are normally covered by a tectorial membrane which has been removed in this specimen. ~ 4 6 8 . From Miller (‘811.

Fig. 6. Frontal section of the left papilla basilaris of Anolis equestris. The larger hair cells located apically are unidirectionally oriented. ~ 2 5 5 .

Fig. 7. Cross section of the papillar nerve of Anolis equestris. While larger nerve fibers occur throughout the nerve, those in the apical unidirectional hair cell region (reader’s left1 are mostly larger in diameter. x405.

Fig. 8. SEM of the left papilla basilaris of the iguanid, Dipsosaurus dorsalis. Note the large extent of the centrally located unidirectional hair cells. x600. From Miller (‘81).

Fig. 9. Frontal section of a right papilla basilaris of Dipsosaurus dorsalis showing the larger unidirectional hair cells in a central location. The spaces around many hair cells (arrows) are expanded regions of the innervating nerve fibers. x 565.

Fig. 10. Cross section of a left papillar nerve of Dzpsosaurus dorsalis. The larger nerve fibers tend to be centrally located. ~ 5 6 0 .

8 M.R. MILLER

Fig. 11. SEM of the tropidurine iguanid, Plicaplica. The apical unidirectional hair cells (reader’s left) are covered by a tectorial membrane. The bidirectional hair cells have longer stereocilia adjacent to the apical region and become progressively shorter toward the basal end. X338.

Fig. 14. Frontal section of the papilla basilaris of the tropidurine iguanid, Leiocephalus schreihersi, showing the larger unidirectional hair cells located in the mid-papillar region. x341.

Fig. 15. Cross section of the papillar nerve of Leiocephalus schreihersi. The larger nerve fibers are not as clearly segregated and related to the unidirectional hair cell region as they are in most species. ~ 5 4 0 .

Fig. 12. Frontal section of a left papilla basilaris of Plica umbra showing the larger unidirectional hair cells located apically. ~ 2 3 8 .

Fig. 13. Cross section of the left papillar nerve of Plica plica. Note the segregation of larger nerve fibers a t the apical end. x338.

Fig. 16. SEM of a left papilla hasilares of the anguid, Celestus costatus, showing a short apical region of unidirectional hair cells. The tectorial membrane has been removed. The stereocilia of the bidirectional hair cells are longest near the apical unidirectional region and become progressively shorter basally. x450.

Fig. 19. SEM of a left papilla hasilaris of Ophisaurus uentralis demonstrating centrally disposed unidirectional hair cells in an anguid. Unidirec- tional hair cells are more often apical in anguids. x494.

Fig. 20. Cross section of the papillar nerve of Ophisaurus uentralis showing a tendency for segregation of larger fibers in the mid-portion of the nerve. ~ 4 5 0 . Fig. 17. Frontal section of the papilla basilaris of Celestus costatus show-

ing the larger unidirectional hair cells in the apical region. ~ 2 5 7 .

Fig. 18. Cross section of a left papillar nerve of Celestus costatus showing very pronounced segregation of larger fibers apically. x 494.

10 M.R. MILLER


moderately large and are unusual in lizards because 85- 95% of the hair cells are abneurally oriented. All the hair cells are covered by a relatively heavy limbus-attached tectorial membrane. While SEM does not show differences in hair cells other than kinocilial orientation, histologic section (Fig. 22) of the papilla shows that a large central portion contains relatively large hair cells, while both terminal regions consist of smaller and shorter hair cells. In a corresponding section of the papillar nerve (Fig. 23) larger nerve fibers supply that region of the papilla containing the larger hair cells. Figure 4 depicts the hair cell orientation of a teiid lizard. This pattern of kinocilial orientation is based on that of the teiid, Tupinambis teguixin, which is much different from that of Ameiva and Cnemidophorus. SO much structural variation in the auditory papillae is unusual between species in lizard families.

Varanidae (Table la). The papilla basilaris of Varanus exanthematicus is very similar to that of I? bengalensis (Miller, '78b) and consists of a group of unidirectionally oriented hair cells located in the apical quarter of the basal segment, and bidirectionally oriented hair cells occupying both the basal portion of the basal segment and the entire apical piece (Fig. 4). Section of the papillar nerve shows larger nerve fibers supplying the portion of the papilla containing the unidirectional hair cells.

Gekkonids flable la, Figs. 24, 25). The length of the papilla basilaris varies markedly in the gekkonid lizards, from ca. 0.7 mm in smaller species to 2.5 mm in larger species (Miller, '66, '73a; Wever, '78). Hair cell number increases with papillar length (the coefficient of correlation (r) for papilla length and hair cell number for 11 species is .846). As shown in Table la , hair cell numbers for the four species used in this study varied from 435 to 2,200. Wever ('78) reported that in Gekko smithi the hair cell number was 2,500, the largest so far reported in a lizard species.

Regardless of papilla length or hair cell number, the structural features of gekkonid papillae are remarkably similar. In all the species studied here the papillae con- sisted of an apical region composed of two longitudinally separated strips of bidirectional hair cells and a basal region of unidirectionally oriented hair cells (Figs. 4,241. The bidirectional hair cells are covered by two types of modified tectorial membranes, and the basal unidirectional hair cells

Fig. 21. SEM of a left papilla basilaris of a teiid lizard, Cnemidophorus motaguae. Higher power observation reveals that only the abneuralmost row of hair cells are neurally oriented. Most of the t, ctorial membrane has been lifted off the hair cells and only a hit of the apical portion remains attached to the hair cells. x 188.

Fig. 22. Sagittal section of the papilla basilaris of Cnemidophorus rnotu- gum showing larger centrally located unidirectional hair cells and small hair cells a t both ends. Most of the smaller hair cells are abneurally oriented, however. x 194.

Fig. 23. Cross section of the papillar nerve of Cnernidophorw motugum showing that the larger nerve fibers are positionally related to the region of the larger hair cells seen in Figure 22. X261.

Fig. 24. SEM of the papilla basilaris of the gekkonid, Coleonyx uuriegu- tus. The unidirectional hair cells are located in the narrower basal region (reader's right). Except for a few apical sallets all the tectorial covering has been removed. x 103. From Miller ('73b).

Fig. 25. Cross section of the papillar nerve of Coleonyx uuriegaius showing larger nerve fibers a t the basal end of the nerve corresponding to the basal unidirectional hair cell regions of the papilla (Fig. 24). X353.

by fine strands of tectorial material (MIller, '73a). In a section of the corresponding papillar nerve (Fig. 25), the larger nerve fibers correspond in position to the basal unidirectional hair cell region. It should be noted that among lizards the basal location of unidirectional hair cells found in the gekkonids (and pygopodids) is unique and occurs only in species of these families. Also significantly, the tonotop- icity of these membranes is reversed in that low frequency reception is associated with the basal unidirectional hair cell region (Manley, '81). In other species, low frequency reception is either apical or mid-papillar depending on the location of the unidirectional hair cells (see discussion).

Scincidae (Table la; Figs. 4, 26-28). The papillae basilares of the species of skinks used in this study (Fig. 26) are typical of other skink species (Fig. 27) (Miller, '74). An apical group of both bi- and unidirectionai hair cetis are covered by a large tectorial structure (the 'culmen', Wever, '781, while the very regularly arranged bidirectional hair cells occupying the curved elongated basal strip are covered by specialized tectorial formations (sallets). A cross section of the papillar nerve (Fig. 28) shows no particular segregation of nerve fiber sizes.

Cordylidae (Table la). Unfortunately only two specimens of a cordylid lizard could be obtained for this study because no other South African lizards were available. A hair cell number estimate was made on the basis of a previous study of Cordylus jonesii (Miller, '78b). Structur- ally the papilla basilaris of the cordylids is very similar to that of the skinks. The papillar nerve does not contain the diversity of fiber size present in the skink and there is no indication that larger nerve cells supply the apical unidirectional hair cells.

Xantusiidae (Table la; Figs. 29-31). The relatively small size and especially the fragility of the papilla basilaris of Xantusia uzgzlis makes hair cell number determination very difficult. Thus the numbers cited for hair cell content in Table l a are estimates. Like that of the cordylids, the papilla of Xantusia uzgilis (Miller, '78b; Figs. 29, 30) is structurally similar to skinks. Section of the papillar nerve (Fig. 31) shows no definite segregation of 1arge-r. nerve fibers corresponding in position to the apical unidirectional hair cells.

Variations in hair cell numbers Data on variation in the numbers of hair cells in 162

specimens of 15 lizard species are given in Table la . The coefficients of variation (standard deviatiodmean x 100) of hair cell numbers for these species varied from 3.2 to 16.6. For strictly qualitative purposes the averages of these coefficients of variation is 7.6. With a coefficient of variation of 7.6 in a population of hair cells in which the mean number is 100, one st,andard deviation (66% of the population) would be f 7.6, or a percentage'variation of 15.2. Two standard deviations representing 96% of a population is then a variation of approximately 30%. The average coefficient of variation for papillar nerve fiber numbers in 381 specimens (35 species) is 7.6, the same average found for hair cell variation. It will be shown later (Discussion) that variations in the numbers of auditory hair cells and nerve fibers in lizards are similar to those found in other vertebrate species.

The difference in hair cell number between the right and Ieft ears of individual animals was determined in three species and the differences expressed as the proportion of variation due to the right-left ear differences. The proper

12 M.R. MILLER

statistical method for deriving this proportion was provided by The Computer Center, University of California, San Francisco.’

Differences in hair cell numbers between the right and left ears of individual animals:

Species Number Proportion of

pairs of due to the papillae R-L ear

difference

of the variation

Crotaphytus wislizeni 5 9.68

Dipsosaurus dorsalis 6 5.5%

Sauromalus obesus 4 9.4%

There is no correlation between hair cell number-and age (age in lizards is proportional to the size of an animal usually expressed as the snout-vent length), sex, or sidedness (right or left ear) in any lizard species.

Nerve fiber numbers That portion of the papillar nerve (Figs. 2, 3) from the

posterior ganglion (which is usually located within the otic capsule) to the papilla basilaris varies in length from ca. 0.2 to 0.75 mm. The nerve fibers do not branch within the nerve. While no amyelinated nerve fibers were observed by light microscopy, or by a few TEM observations, an insufficient number of specimens were studied by TEM to confirm their absence or presence. In the lizards studied, the number of hair cells varied from ca. 50 to 2,000 and the number of nerve fibers from ca. 300 to 2,000.

‘Method of calculating the proportion of overall variability ac-

Variation between left and right ears as a percentage of the total counted for by differences between ears.

variation from measurements on both ears of multiple animals:

Let Xij =measurement on animal i, ear j, for i = 1, ----, n; j = 1, 2 (1 = left, 2 = right).

2

Let Y = C Xij == sum for animal i. J - 1

n 2

Let Z = C C Xij = overall sum. 1 - 1 J - 1

Compute corrected total sum of squares (SSTO)

sum of sauares for blocks (animals) (SSBL)

% variation between ears relative to total

SSTO - SSBL SSTO

= 100 x

= “proportion of overall variability accounted for by differences between ears.”

The coefficients of variation for the number of nerve fibers in 35 species ranged from 1.2 to 14.4 with an average variation of 7.6.

When both the hair cell and nerve fiber numbers were determined in the same animal, the correlation between hair cell and nerve fiber number was as follows:

Species

Crotaphytus collaris baileyi

Crotaphytus collaris collaris

Crotaphytus wislizeni

Sceloporus magister

Celestus costatus

Number of

specimens 5

5

17

11

6

dcoefficient of correlation)

0.89

0.46

0.88

0.25

0.87

The difference in the number of nerve fibers between the right and left ears of individual animals (108 pairs) from 24 species (eight families) of lizards is shown in Table 2. This table demonstrates that the proportion of variation in nerve fiber numbers due to right-left ear differences ranged from 2 to 11%.

Nerve fiber number is not correlated with age (size), sex, or sidedness (right or left ear).

Nerve fiber number is not directly related either to papilla basilaris length nor to species size since different structural types of papillae have different papillar nerve fiber numberhair cell number (PNFNMCN) ratios, and it is these ratios rather than the numbers of hair cells or nerve fibers which have significant functional and phylogenetic relationships.

Papillar nerve fiber number hair cell number ratio (Table lb)

Figure 32 is a graph demonstrating the relationship between papillar nerve fiber and hair cell number (PNFN/ HCN ratio) in ten lizard families. The four iguanid sub- groups (anolines, iguanines, sceloporines, and tropidurines) have the largest ratios (4.2-7.4). Species of the Agamidae and Anguidae have slightly smaller raitios (3.7-3.8) than the Iguanidae. All species of the above three families have papillae basilares with apically or centrally located tectorial-membrane-covered unidirectional hair cells and basal or terminal groups of bidirectional hair cells without a tectorial membrane. Figure 33 demonstrates the relationship between the PNFN/HCN ratio and species adult body size and shows that in the iguanids, agamids, and anguids there is a tendency for larger species to have lower PNFN/ HCN ratios. The coefficient of correlation between the PNFN/HCN ratio and body size for the 18 species of iguanids listed in Fig. 23 is -.548. The negative correlation indi- cates that the PNFN/HCN ratio decreases as body size increases. Species of the above families with lower ratios also have longer papillae and larger numbers of hair cells.

While all hair cells in species of the family Lacertidae are short-ciliated and superficially alike as seen by SEM and are all covered by a tectorial membrane, the PNFNMCN


Fig. 26. SEM of a right papilla basilaris of the scincid lizard, Eumeces luticeps. The apical hair cells (reader’s left) are covered by a large tectorial mass. The bidirectional hair cells extending basally from the apical tip are covered by specialized tectorial formations called sallets. x 86.

Fig. 27. SEM of the left papilla basilaris of the skink, Mabuyu carinata, showing the hair cells with the tectorial membranes removed. A few hair cells at the tip of the apical expansion are the only unidirectional hair cells in ski* papillae. x62. From Miller (‘74).

Fig. 28. Cross section of the papillar nerve of Mubuyu parrotetii. There is no particular segregation of nerve fiber sizes in skink papillar nerves. x 370.

13

Fig. 29. SEM of the basal portion of the papilla basilaris of Xantusk uigilis. All the hair cells in this region are bidirectional and are covered by specialized membranes (sallets) which have been removed in this specimen. x295. From Miller (‘78b).

Fig. 30. SEM of the entire papilla basilaris of Xantusia uigilis showing the enlarged apical expansion (arrow) similar to that of skinks. x48. From Miller (‘78b).

Fig. 31. Cross section of the papillar nerve of Xuntusia uigilis. While there is less mixture of varying nerve fiber sizes than in skinks (Fig. 28), there is no definite segregation of larger fibers toward the apical end of the nerve. ~ 6 0 8 .

14 M.R. MILLER TABLE 2. Differences in Nerve Fiber Numbers Between Right and Left Ears -

Proportion of No. Range of PNFN' Range of differences variability due to of in all paired between right and right-left ear

differences2 (70, - Species pairs specimens left ears

I gu a n i d a e (Anolines)

Anolis carolinensis Polychrus marmoratus

Crotaphytus wislizeni C. collaris baileyi C. collaris collaris Dipsosaurus dorsalis Sauromalus obesus

(Sceloporines) Uta stansburiana Scelnporus magister

(Tropidurines) Tropidurus hispidis Plicaplica Plica umbra Uranoscodon supraciliaris


Celestus costatus Ophisaurus ventralis

Lacerta sicula

Ameiva ameiua Cnemidophorous tigris

Coleonyx uariegatus Gekko gecko Hemidactylus frenatus

Mabuya parrotetii

Qguanines)

Agamidae

An gu i d a e

Lacertidae

Teiidae

Gekkonidae

Scincidae

Xantusiidae

4 3

9 3 5 7 5

5 5

5 4 3 3

3 3

5 3

4

3 6

4 4 3

3

775-924 420-534

560-752 1,150-1,288

854-1,244 407-570 468-616

374-460 550-647

1,246-1,516 1,521-1,693 1,522-1,782

99 1-1, I43

1,064-1.159 1,050-1,286

1,240-1.634 693-884

479-601

1,654-1,823 998-1,236

581-760 1,667-2,102

489-635

928-1,082

3-35 9-20

6-78 2-59

35-68 4-31 1-23

5-26 5-48

11-45 18-54 45-72

1-35

9-25 7-43

4-82 8-34

6-31

1-29 8-62

5-43 14-55 6-25

14-38

4.26 3.45

7.15 9.88 5.68 3.27 2.03

6.82 10.55

3.17 10.33 9.68 4.79

7.98 2.28

3.13 2.62

9.09

2.02 11.00

6.10 4.22 2.99

5.26

Xantusia vigilis 6 355-433 9-20 5.79

'Papillar nerve fiber No. 'See text for method of derivation

ratio is relatively high (5.3). However, more species of this family should be studied to more accurately assess the status of the PNFNMCN ratio.

In teiid species, PNFN/HCN ratios vary from 2.4 to 3.2 (Table lb, Figs. 32, 33). In the listed species all hair cells are covered by an unmodified tectorial membrane, and a large proportion of the hair cells are abneurally oriented. Figure 33 shows no correlation between the PNFNMCN ratio and the adult body size (snout-vent length) of species.

Unfortunately only one varanid species was studied, and its PNFNMCN ratio was 1.1, which is similar to that of other species with longer papillae containing more numer- ous hair cells (vide infra). The papillar structure of Varanus is less complex than that of gekkonids and scincids in that all hair cells are covered by an unmodified tectorial membrane, and the unidirectional hair cells are centrally and the bidirectional hair cells apically and basally located.

In species of gekkonids, scincids, cordylids, and xantusiids, the PNFNHCN ratios ranged from 0.8 to 1.5 (Table lb). In species of these families the papillae are longer with larger numbers of hair cells covered by specialized types of tectorial membranes. Three different types of tectorial cov- erings occur in a single gekkonid species (Miller, '73a). Figure 33 shows no correlation between adult body size and the PNFNBCN ratio in species of these families. There was also no correlation between papillar length or hair cell number and the PNFNBCN ratio.

It is important to note the variability in the PNFN/HCN ratio within species (Table 4). The significance of the varia-

bility in the ratio applies particularly to those species with a high ratio such as Crotaphytus wislizeni, where individual hair cells may be supplied by an average number of nerve fibers ranging from 9.6 to 13.3. Ik is probable that had larger sample sizes been available in several of the other species studied, the overall extent of variation in the PNFN/HCN ratio would have been greater.

Composition of the papillar nerve (Table 3). Probability plots of several papillar nerves demonstrated clearly that the contained nerve fibers were unimodally distributed. More than 99% of the nerve fibers ranged in diameter from 0.8 to 6.0 pm in 35 species. Figures 34-36 are histograms of the mean percentages and standard deviations of papillar nerve fiber diameters in 18 of the species studied.

In a few iguanid, agamid, and anguid species (Phryno- soma platyrhinus, Acanthosaura crucigeera, and Ophisaurus ventralis) there is no clear correlation between papilla basilaris structure and papillar nerve fiber distribution. In those species with a small percent of apical unidirectional hair cells (anolines and some tropidurines) or with only two rows of bidirectional hair cells (sceloporines), the larger percent of nerve fibers were less than 3 pm in diameter (0.8-3 pm; Fig. 34). In species having rlelatively large per- cents of unidirectional hair cells (iguanine iguanids) or lower PNFNMCN ratio (teiids, varanids, scincids, gekkonids, cordylids, and xantusiids), more than half the nerve fibers were larger than 3 pm (3-6 pm; Figs. 35,36).

Calculations were made to determine the area of the nerve fibers in each 1-pm group of nerve fibers. The infor-


1800

1600

1400

1200

1000

800

600

400

200-

15

1 /1 lguanines 7 3

lguanidoe Sceloporines 7 4

Tropidurines 5 0 Agomidae 3 8

Anguidae 3 7 / Varanidoe

Teiidae 2 6 Varonidae 1 1

kincidae 0 8

Xantusiidae 1 1

/ /

Lacertidae 5 3 m

Gekkonldae 0 9 /

/

- /’ -

/ /

- - Cordylidoe 0 8

- /’ - /

/ /

- /

8 / Scincidae 8

- Gekkonidae

/

- /’ /

/ - Cordylidoem / /

/ Anguidoe

/

Agomidoe8

- Xantusiidae8

(1)Tropidurines / // (1)Anolines

- / Locertidoe8

/ (Weloporines. ‘(1)lguanines I I I 1 I l I I l ~ l l l ~ l l l

200 400 600 1000 1200 1400 1600 1800

Relationship Between Papillar Nerve Fiber Number and Hair Cell Number (PNFN/HCN) in Lizard Families

Teiidae

Mean Papillar Nerve Fiber Number

Fig. 32. Graph demonstrating the relationship between papillar nerve fiber number and hair cell number in lizard families. The black squares represent the means of the papillar nerve fibers and the hair cell numbers for each family. The dashed line which is a t a 45” angle to the X and Y axes represents points where a ratio of lil would be located. Ratios less than l i l would be above this line, and ratios greater than lil, below this line

mation derived from comparison of nerve fiber areas in these size groups in different species did not add any information to that provided by the data (percent of nerve fiber sizes in each 1-pm group of fibers) given in Table 3.

Serial sections of the basilar papilla showed that larger nerve fibers supplied unidirectionally oriented hair cells (see hair cells). It was also observed that the larger the area of unidirectional hair cells, the greater the number of larger nerve fibers.

For the purpose of comparing the nerve fiber content of an auditory nerve with that of a vestibular type, nerve fiber numbers and sizes were determined in the nerve to the posterior ampulla (Fig. 2) in one specimen of each of ten species from seven families of lizards. Ampullar nerve fiber numbers varied from 389 to 1,214 and scatter plots revealed a definite bimodal distribution, with one group of fibers 1- 5 pm and another group 5-14 pm in diameter. The number of smaller nerve fibers ranged from 50 to 90% of the total number of fibers. Studies of nerve fibers innervating the

hair cells of the crista ampullaris showed that the larger nerve fibers supplied the calyceal endings associated with type 1 hair cells and smaller nerve fibers supplied type 2 hair cells. A similar pattern of innervation was observed in the macula lagenae.

Detailed pattern of hair cell innervation. The complete pattern of hair cell innervation in the iguanid lizard, Sce- loporus occidentalis, has been determined in my laboratory by Teresi (’84). Serial TEM sections of an entire auditory papilla revealed that all nerve fibers innervate only one hair cell. The unidirectional hair cells are supplied by four to 17 afferent nerve fibers (mean of ten) and 0 to five efferent nerve fibers (mean of 2.7). Bidirectional hair cells are supplied by five to 13 afferent fibers (mean of ten) but do not receive any efferent innervation.

The presence of amyelinated nerve fibers in the papillar nerve has been commented upon earlier. We have not as yet determined the number of efferent nerve fibers in the papillar nerve, but extrapolating from Teresi’s (‘84) studies,

16 M.R. MILLER

TABLE 3. Papillar Nerve Fiber Sizes (Mean Percent and Standard Deviation of Papillar Nerve Fibers in 1-pm Increments)

No of 0-1 um 1-2um 2-3urn- 3-4um 4 5um

measured M%' SD? M% SD Ma SD M'ic SD M% SI) specimens

~

S~ecies

Iguanidae (Anolines)

Anolis carolinensis Anolis equestrcs Polychrus marmoratus

(Iguanines) Crotaphytus wislczeni C collaris baileyi C. collaris collaris Dipsosaurus dorsalis Phrynosoma platyrhin u s Sauromalus ohrsus

(Sceloporines) Callisaurus draronoidcs Uta stanshurianu Sceloporus magister

Clkopidurines) Tropidurus hispidis Plica plica (a)' Plica plica m, Plica umbra Uranoscodon supraccliaris Leiocephalus schreihersi


Celestus costatus Ophcsaurus uentralis

Lacerta sicula ( a ~ 4 Lacerta sicula (b)

Ameiua ameiua Cnemidophorus motaguae Cnemidophorus tigris Callopistes maculata

Varanrs exanthematicus

Coleonyx uariegatus Cosymhotus platyurus Gehho gecho Hemidactylus frcnatus

Eumeces laticeps Eumeces ohsoletus Mahuya multifasciata Mahuyu parrotetii

Cordylis ucttifer

Xantusia vigilis

A g a m i d a e

Anpidae

Lacertidae

Teiidae

Varanidae

Gekkonidae

Scincidae

Cordylidae

Xantusiidae

~

8 4 5

9 6

4 5

-

-

5 10 4

5 4 4 5 5 6

7 6

5 6

4 4

5 6 6 2

3

8 5 7 7

5 5 4 5

2

.05 .05 44.0 12.3 42.9 8.8 12.4 5.3 0.6 0.6

.09 .07 16.0 2.8 46.8 11.1 33.7 10.3 3.2 2.5 - - 27.4 7.5 45.4 5.1 23.8 5.4 3.3 1.0

- - 6.8 1.5 36.3 7.1 48.6 5.0 8.1 4.3 - - 4.5 3.1 23.8 5.0 53.3 5.9 17.5 2.1

- - 7.2 .8 25.5 3.6 51.6 3.8 15.0 7.0 .04 .09 12.2 4.0 34.2 7.4 47.2 4.0 6.1 3.9

- - 15.9 5.6 52.7 8.2 30.4 10.2 1.0 0.5 .14 .25 26.5 7.8 41.7 8.7 29.0 5.6 2.6 2.3 .12 .15 23.5 2.9 54.1 5.4 20.6 2.7 1.7 0.8

.08 .I0 26.5 4.3 45.0 3.7 25.7 1.9 2.8 1.0

.08 .03 10.7 1.4 27.6 4.9 45.5 1.3 14.6 4.1

.16 .09 25.2 7.2 47.1 2.8 24.5 8.4 3.0 1.3 6 6 6 0 50.8 6.5 36.0 5.0 11.6 1.7 0.9 0.3 .04 .08 5.8 2.8 14.3 1.9 46.9 4.2 28.4 2.5 .05 .06 13.8 3.9 33.7 5.3 41.1 6.4 10.8 4.6

.05 .07 6.2 3.1 27.9 9.2 51.4 6.6 12.9 6.4

.01 .03 18.6 7.4 44.8 2.6 34.0 7.9 2.6 1.6

.44 .45 28.3 8.6 55.3 5.6 14.1 6.7 1.6 .6 - - 9.7 2.9 39.1 9.3 44.1 7.0 6.8 4.8

- - 2.7 0.2 16.2 3.3 55.4 2.7 24.6 2.8 -- - 9.0 1.2 39.5 7.2 47.7 5.5 3.7 2.7

.O1 .02 2.2 1.0 12.2 2.8 46.3 4.0 34.5 4.3

.03 .06 7.7 2.9 27.0 6.0 42.5 5.4 18.5 4.8

.02 .04 8.3 2.1 29.7 3.4 44.0 4.1 17.2 4.4 0.9 - 2.5 - 21.3 - 58.6 - - -

- 2.6 0.4 10.2 1.8 43.1 4.1 38.5 4.5

- - 4.9 1.7 23.0 5.5 50.7 5.1 18.0 7.6 - - 6.6 2.9 22.6 5.4 49.8 5.6 18.0 6.1 .06 .05 3.4 1.3 15.1 4.5 51.2 4.6 25.8 6.5 .03 .08 7.7 1.9 30.5 4.5 52.5 4.0 8.6 3.0

- - 5.2 2.0 33.4 6.9 49.0 6.5 10.4 3.6 .16 .22 4.0 2.4 27.3 11.7 47.7 3.8 17.3 11.9 .03 .05 13.4 1.5 37.3 6.6 36.9 1.7 10.9 1.7

6.9 2.6 28.5 1.9 35.4 4.7 17.8 1.4

6.9 - 18.2 - 49.4 - 22.0 -

- -

- -

M 9 SD

.03 .07

.07 .05

.05 .09

.26 .49

.97 .59

.68 .41

.12 .I9 - -

.09 .09

.04 .09

.08 .09

- -

1.4 2 5 .06 .05 .04 .09

.53 .70

1.4 .77 .06 .05

2 4 .34 2 2 .15

1.2 38 .08 .10

4.6 .SO 3.8 1.9

.67 .31 14.8 -

3.9 1.3

5.1 .70

2.9 1.8 2.5 2.2 3.9 2.6 51 .50

1.6 1.5 3.3 3.3 1.5 1.8 8.5 2.0

3.5 -

-

.03 -

-

.02

- - -

- - -

-

.08 - -

0.18 -

.05 -

-

0

.05 -

-

.18

.02 2.0

2 7

.16

.20

.19 3 0

.18

.il .08 .67

0

31

87.0 62.9 72.8

43.1 28.3

32.7 46.4 -

68.6 68.3 77.7

71.6 38.4 72 5 87.5 20.1 47.6

34.2 63.4

84.0 48.8

18.9 48.5

14.4 34.7 38.0

3.4

12.8

27.9 29.2 18.7 38.2

38.6 31.5 50.7 35.4

25.1

34.2

13.0 37.0 27.2

57.0 71.8

67.3 53.4 -

31.5 31.6 22.4

28.5 61.6 27.6 12.5 80.0 52.4

65.8 36.7

15.9 51.1

81.3 51.5

85.6 65.4 61.9 96.6

87.2

71.9 70.7 81.5 61.9

61.5 68.6 49.4 64.4

74.9

65 8 11 .02 .06 7.4 3.0 26.8 9.2 40.7 4.4 19.4 7.2 4.7 1.9

3.8 15.0 29.6

50.0 47.0

51.0 40.0 38.0

38.0 25.0 28.0

16.0 13.0 13.0 22.0 16.0 25.0

17.0 28.0

11.0 20.0

11.0 11.0

31.6 35.0 67.0

Unknown

13.0

13.05 10.05 10.05 10.05

4.05 4.05 4.05 4.05

20.0

17.0

'Percent of unidircctionally oriented hair cells. %an percent "Standard deviation. 'Apparently two different populations. 'Estimated


1 80 h

E E

TABLE 4. Range in the PNFN/HCN' Ratio in Split Specimens'

m G e k k o gecko @ Gotaphytus mllwrs collaris - @ Crotofiytus wrslirenr - @I Dipsosourus dorsalis

@ Plica prlco

Species

5 - - 100-

E t 8

3 -0 0 v

80 iii

.- 6 0 - Q v)

40

PNFN/HCN Ratios ~ -

PNFN HCN No. of - SD CV specimens M3 SD4 CV5 M SD CV Range M

la 0 0

03

0 0

@ @

a @ @ Q @ -

0 0

I I ~ I ~ I I I I I ~ I I I ~ I ~ I I ~ I I

Crotaphytus wislizeni 18 656.6 80.0 12.2 61.8 7.8 12.6 9.6- 10.7 .9 8.5

Crotaphytus collaris 13.3

haileyi 4 1,224 76.3 6.2 121 11.6 9.6 9.6- 10.2 .5 4.8

Crotaphytus collaris

Dipsosaurus dorsalis 20 499.4 53.9 10.8

Sceloporus magister 10 612.7 44.9 7.3

Lewcephalus schreihersi 3 793.7 8.3 1.1 Plica plica 6 1,582.8 68.4 4.3 Plica umbra 4 1,645.3 121.4 7.4

collaris 6 979.5 78.9 8.1

Sauromalus obesus 11 531.4 70.4 13.2

Agama agama 4 1,121 31.4 2.8

Celestus costatus 6 1,554 49.7 3.2

'Papillar nerve fiber No.iHair cell No. 'Specimens in which both the PNFN and HCN were determined in the same specimen. %an. 4Standard deviation.

'~oefficient of variation = :X 100 SD

M

114.7 7.7 6.7 89.5 5.4 6.0

139.6 7.9 5.6 87.4 3.9 4.5

294.8 14.0 4.7 116.3 10.8 9.3 346.8 38.4 11.1 333.5 28.6 8.6 398.2 21.8 5.5

10.7

6.8-9.6 4.3-6.4 3.2-5.3 6.4-8.8 3.7-3.9 6.4-7.6 3.9-5.2 4.8-5.0 3.7-4.0

8.6 5.6 3.8 7.0 3.8 6.9 4.6 4.9 3.9

1.0 .5 .6 .I .1 .I .5 .1 .1

12.1 9.7

16.6 10.4 2.2 9.6 9.7 2.2 3.1

Relationship Between Papillar Nerve Fiber Number/ Hair Cell Number Ratio and Adult Body Size of Different Species

Fig. 33. Graph demonstrating the relationship between the papillar nerve fiber number and hair cell number ratio and the adult body size of different lizard species. Note that the ratio decreases as body size increases in the iguanid-agamid-anguid species (r = -.548) but not in the teiids, gekkonids, or scincids.

Anolis carolinensis a 6ol

6 C

5c

40

30

20

10

%

60

50

40

30

20

10

%

Tropidurus hispidie C

p m 1 2 3 4 5 6

Ceiestus costatus e T

p m 1 2 3 4 5 6

Fig. 34. Histograms of the distribution of nerve fiber sizes in the papillar nerves of species in which the larger percentage of nerve fibers is 0.8-3 pm in diameter. Of the six species shown here, Celestus costatus (e) is an anguid and all the others are iguanids. The papillae basilares of these species are

approximately 25% of the nerve fibers supplying the unidirectional hair cell regions might be efferent fibers.

DISCUSSION Hair cell number

The auditory hair cells of fish are not considered here because the markedly different structure of the auditory portions of the fish lagena, utriculus, and sacculus are not comparable to the papillae basilaris of reptiles. The papilla amphibiorum of amphibians is also not included because it is not homologous with the reptilian auditory papilla. The

60

5 0

40

30

20

10

%

60

50

40

30

20

10

%

60

5 0

40

30

20

10

%

h 78%

Sceloporus mapister

b

p m i 2 3 4 5 6

Acanthosaurs crucipers

d

p m l 2 3 4 5 6

Plica umbra f

m p m 1 2 3 4 5 6

characterized by relatively small percentages of unidirectional hair cells and large numbers of bidirectional hair cells. In Figures 34-36 the figures within the clear bars represent the total percent of nerve fibers 0.8-3 Wm in diameter. Ordinate, percent of nerve fibers; abscissa, nerve fiber diameters.

data in Table 5 cited for hair cell and nerve fiber numbers in the papillae basilares of Ambystoma tigrinum and Rana catesbeiana show a larger range in hair cell numbers but a range in nerve fiber numbers comparable to that of lizards.

Wever (‘78) gives hair cell numbers for a large variety of lizards. These data are based on small isample sizes, however. While Wever’s and my data are in the same range when reporting hair cell numbers of the same species, Wev- er’s figures are usually ca. 10% less than mine. These differences could be attributed to studying different populations of a species, but probably are due to the use of different methods to derive hair cell numbers. It is probable


AQSma sgsma a 6o I T

19

60

50

40

30

20

10

%

60

50

40

30

20

10

%

p m i 2 3 4 5 6

Leiocephalus schreibersi C

i m 1 2 3 4 5 6

Dipsosaurus dorsalia e T

p m 1 2 3 4 5 6

Fig. 35. Histograms of the distribution of nerve fiber sizes in three species of iguanids (c-e), an agamid (a), and a lacertid (0. In all these species, more than half of the nerve fibers are greater than 3 pm in diameter (3-6 pm). In b-e, larger nerve fiber sizes may be correlated with a relatively

that using a combination of SEM and serial frontal sections of the papillae to count hair cells as was done in this study may provide a more accurate result than Wever's method of counting hair cell nuclei in serial sections of papillae.

Table 5 gives the hair cell and nerve fiber numbers for two lizard species. It is interesting that Gerrhonotus multicarinatus and Celestus costatus (reported in this study), both anguids, have similar PNFNLHCN ratio (4.1 for Gerrhone tus and 3.7 for Celestus). Calotes uersicolor, an agamid, has a PNFNkICN ratio of 3.6, similar to that of the agamid, Agama agama (3.7) reported in this study.

Ophlsaurus ventrslis b

50

4 0

30

20

10

'16

60

50

4 0

30

20

10

96

60

50

4 0

30

20

10

x

Crotaphytus wislizeni d

p m 1 2 3 4 5 6

Lacerts sicula (b) f

T

i r n 1 2 3 4 5 6

large number of unidirectional hair cells. Why a and f that do not have larger numbers of unidirectional hair cells fall into this group is unex- plained. Ordinate, percent of nerve fibers; abscissa, nerve fiber diameters.

Considering that the length of the papillae basilares of Caiman crocodilus and Alligator mississippiensis are each approximately 4 mm (personal observations), the reported hair cell numbers are large (11,500 and 11,000, respec- tively). Such a large number of hair cells in a relatively short papilla is due to the width of the crocodilian papilla (20-30 rows of hair cells). The structure of the pigeon papilla is similar in this regard (Smith, '81). The PNFNLHCN ratio of the pigeon calculated from the data in Table 5 is approximately 0.5, which is the lowest ratio so far reported in any vertebrate species.

20 M.R. MILLER

80

50

4 0

30

20

10

%

60

50

40

30

20

10

%

60

50

40

30

20

10

x

Arnelva arnelva a

Coleonyx variegatus

C T

u r n 1 2 3 4 5 8

Mabuya parrotetii e

p r n l 2 3 4 5 6

Fig. 36. Histograms of the distribution of nerve fiber sizes in a teiid !a), varanid (b), gekkonid (c), two scincids !d,e), and a xantusiid (0. In all these species the preponderance of larger nerve fibers (3-6 pm) is correlated with low PNFNiHCN ratios. It is probable that larger nerve fibers are necessary

In mammals hair cell number is related t o papilla length (Lewis et al., ’84) but not as closely as in lizards. The only report of the variation in hair cell numbers in a mammal is that of Bredberg (‘68) for ten human fetal cochleae. Since this report provided no statistical data, only an estimate of the percentage variation may be made by dividing the range by the mean number of hair cells, thus giving an estimated variation of 34%, a figure not far from the percentage variation in lizards (30.4%).

60

50

40

30

20

10

%

80

50

40

30

20

10

%

80

50

40

30

20

10

%

Varanua exanthernatIcu8

b

p m 1 2 3 4 5 8

Eumeces laticeps d

p i 2 3 4 5 6

Xantusia vigilis f

p r n l 2 3 4 5 6

to innervate a larger number of hair cells as compared with species with high PNFNIHCN ratios where nerve fibers mainly supply one or, at the most, a few hair cells. Ordinate, percent of nerve fibers; abscissa, nerve fiber diameters.

Nerve fiber number

From Dunn’s (‘78; Table 5 ) study of the nerve to the papilla basilaris of the bullfrog (Ram catesbeiana) one may calculate a coefficient of variation of 5.2. The larger variation in lizards (7.6) is possibly a reflection of the much larger number of species and specimens of lizards studied.

There is more information on the variation in the numbers of auditory nerve fibers than in hair cells in mammals


TABLE 5. Numbers of Auditory Nerve Fibers and Hair Cells in Some Sub- Mammalian Vertebrates

Papillary nerve fibers Hair cells

Amphibians Rana catesbeiana 747 * 39 50-95

Ambystoma tigrinurn 30-40 40-80 (Dunn, '78) (Li and Lewis, "74)

(White, '78) Reptiles

Gwrhonotus multicarinatus 617 150

Calotes uersicolor 805 225

Caiman crocodilus - 11,500

(Weiss et al., '76)

(Bagger-Sjoback, '76)

(Leake, '77)

(Wever, '78)

(Wever, '78)

Alligator mississippiensis - 11,000

Crocodilus acutus - 13,700

Birds Pigeon 5,136 10,400

(Boord, '69; Goodley and Boord. '66)

(Table 6). So far the best information is that given by Ehret ('79) reporting the mean and standard deviation for the cochlear nerves of six house mice. Using Ehret's figures for the mean and standard deviation, the coefficient of variation in the mouse is 6.5, which compares well with 7.6 for lizards. Bohne et al. ('82) reported the range and average nerve fiber numbers in eight chinchilla cochleae (Table 6). If one divides the range by the mean nerve fiber number,

there is a variation of ca. 26%, which compares well with 30% found in lizards. The data for the guinea pig, cat, and macaque are based on too few specimens to be useful for comparative purposes. Rasmussen's ('40) report of the nerve fiber numbers of 40 human cochlear nerves, while excellent from the standpoint of sample size, is lacking statistical information. Thus one may obtain only a very rough estimate of the variation by dividing the range by the mean (see Table 6). The variation attained in this manner is approximately 55%, a value considerably larger than that reported for the frog, lizards, or mouse.

While not directly counting the numbers of auditory hair cells or nerve fibers, Hardy ('381, using a reconstructive technic, determined the range and mean length of the auditory papilla in 68 human temporal bones. She reported a mean papillar length of 31.5 mm and a range of 25.3-35.5 mm. The range divided by the mean would indicate a variation of approximately 33%. She also found no significant difference between the right and left ears of the same individual nor any correlation with sex or race.

Nerve fiber sizes in the auditory nerve

Reports on nerve fiber sizes in the auditory nerves of vertebrate species state that the nerve fibers are unimodally distributed and that the largest percentage of fibers are 2-4 pm in diameter (Table 7). The pigeon is exceptional in having the largest percentage of nerve fibers in the 1-2 pm range (Boord, '69). Mammalian auditory nerves also have a component of larger nerve fibers (5-9 pm) not reported in other vertebrates. The percentage of these larger fibers is not great, however.

TABLE 6. Numbers of Auditory Nerve Fibers and Hair Cells in Some Mammals

Auditorv nerve fibers Hair cells

Guinea pig

Cat

Macaque

Chinchilla

House mouse

Harp seal

Bottlenose dolphin

Little brown bat

Man

24,011 (Gacek and Rasmussen, '61)



23,554 (Boord and Rasmussen, '58) 24,703-31,769 Average = 27,123 (Bohne et al., '82)

12,578 i! 819 (Ehret, '79) Coefficient of variation = 6.5

52,000 (Ramprashad, '76)

95,000 (Wever et al., '71)

52,000 (Ramprashad et al., '78)

Average 31,400 (40 cochlear nerves) (range 22,800-40,000) (difference between right and left ears = 4,700) (Rasmussen. '40)

12,500 (Retzius, 1884)

9,300 (Bohne, '76)

3,021 (Ehret and Frankenreiter, '77)

18,000 (Wever et al., '71)

17,400

3.500

Average 16,800 (10 fetal cochleae) (range 14,600-20,400) (Bredberg, '68)

M.R. MILLER 22

TABLE 7. Nerve Fiber Sizes in the Auditory Nerves of Some Vertebrate Species

Nerve fiber No. Fiber diameters

Amphibians Ambystoma tigrinurn

Rana catesbeiana (White, ’78)

(Dunn, ’78) Reptiles

(No reports other than the present work)

Pigeon Birds

(Boord, ’69)

Mammals Mouse

Cat (Ehret, ’791

(Arnesen and Osen, ’78) (Gacek and Rasmussen, ’61)

Guinea Pig

Chinchilla

Monkey

Human

(Gacek and Rasmussen, ’61)

(Boord and Rasmussen, ’58)

(Gacek and Rasmussen, ’61)

(Engstrom and Rexed, ’40)

30-40 25%> J-3 pm 75% 3-8 pm

747 97%’ 0-5pm (myelinated)

Mostly 2-4 pm

5,136 1% 0-1 pm 77% 1-2 pm 21% 2-3 pm 1% 3-4 pm

12.578 Mean = 3.7 pm

1-7 pm (mostly 2-4 pm) 57% 1-3.8 pm

43% 3.8-8.0 pm 78% 1-3.8 pm 22% 3.8-8 pm

1-6 pm (95‘X 2-4 pm) 63% 1-3.8 pm 37% 3.8-8 pm 8.7% 0-2 pm 77.1% 2-5 pm 14.1% 5-9 pm

Difference in the innervation of the right and left ears of the same individual

Reports dealing with the difference between the number of nerve fibers innervating the right and left ears of individual animals are rare and the data inadequate for comparison with my studies. Rasmussen (’40), in his study of 40 human auditory nerves, reported a mean difference of 4,700 fibers between the ears of individuals. A rough estimate of variation obtained by dividing the right-left ear differences by the mean (31,400) would be 15%, a value approximately twice that found in lizards.

Mode of hair cell innervation The precise mode of innervation of the auditory papilla is

the ultimate goal of our present research, and three basi- cally different types of papillae are being studied. Teresi (’84 and unpublished observations) has determined the complete innervation of all 65 hair cells in a specimen of the iguanid lizard, Sceloporus occidentalis, by serial TEM and reconstruction. This is an example of an iguanid-agamid- anguid type papilla with a high PNFNMCN ratio. Each hair cell in this species is innervated “exclusively” by several nerve fibers, i.e., each nerve fiber supplies only one hair cell. Bagger-Sjoback’s study (’76) of the agamid lizard, Calotes uersicolor, is insufficiently complete to state the exact mode of hair cell innervation. In the longer and more complex papillae of the scincids and gekkonids that have low PNFNMCN ratio (0.6-1.5/1), it appears that individual nerve fibers innervate several hair cells (Miller and Beck, unpublished observations). The papilla of the turtle, Chry- semys scripta, an example of a probably primitive type of papilla (Miller, ’78a1, has a low PNFNMCN ratio (1.5) and different modes of hair cell innervation (Sneary, ’84, and unpublished observations). The hair cells overlying the basilar membrane are “exclusively” innervated, while the hair

cells resting on the solid limbic tissue at each end of the papilla are innervated by nerve fibers that are innervating several hair cells (Sneary, unpublished observations).

In Caiman crocodilus, von Diiring (‘74) reported that a single hair cell may make synaptic contact with four afTer- ent neurons, and that up to three hair cells may make contact with the same afferent nerve fiber.

In mammals it is well known that the inner hair cells of the organ of Corti are innervated exclusively by several nerve fibers, while the outer hair cells are innervated by nerve fibers which contact several hair cells.

Thus the mode of innervation found in the iguanid, Sce- loporus occidentalis, is like that of the inner hair cells of mammals. The hair cells of the Caiman may be innervated in a mixed fashion having the characteristics of both the inner and outer hair cells of mammals. The hair cells of the more complex papillae of the scincids and gekkonids may be innervated more in the fashion of the mammalian outer hair cells, but detailed studies of these papillae are still in progress. As more details of hair cell innervation are revealed in future studies the fabric of the innervational picture of the vertebrate auditory papilla will come more into focus and hopefully reveal a sequence that is related developmentally, structurally, and functionally.

Relation of papillar structure to function (Table 8) Increase in the range of frequency reception is related to

the length and complexity of the papilla basilaris in lizards. In species with shorter papillae and higher PNFNMCN ratios (3.5-11.1) as in the iguanid-agamid-anguid type, the upper limit of hearing is 4.3 kHz. In species with longer papillae, larger numbers of hair cells, and low PNFNMCN ratios (0.6-1.5) as in the scincid-gekkonid types, frequencies have been recorded at 5-6.3 kHz. The relationship of sensitivity to either morphological structure or to differences in innervational pattern has not yet been determined.

In all lizard species so far studied lower frequency reception (less than one kHz) is confined to regions of unidirectional hair cells, all of which are always covered by a tectorial membrane. Frequency reception greater than 1 kHz has been found in regions of bidirectionally oriented hair cells. In the shorter lguanid-agamid-anguid type papillae, these hair cells lack tectorial cover. In the longer and more complex papillae the bidirectional hair cells are covered by several specialized types of tecitorial membranes.

Phylogenetic considerations In the last quarter century it has been assumed that the

papillae basilares found in species of iguanid and agamid lizards are “primitive” because these lizards were gener- ally regarded as primitive and the papillae were both short and contained relatively few hair cells (Baird, ’74; Miller ’80). Species of lacertids, teiids, and varanids had larger papillae and were considered “more advanced.” And species of scincids and gekkonids, having the largest papillae, largest numbers of hair cells, and several unusual tectorial modifications were considered to have the “most advanced” papillae.

However, I now believe there is evidence for viewing phylogenetic development of the lizard papilla basilaris in a different perspective. The following observations are the bases for this view.

(1) The Tuatara Sphenodon punctatus and Chelonians (turtles and tortoises) are of more ancient lineage than other living reptiles Otomer, ’661, and it is probable that


TABLE 8. Summary of Functional Data of Some Lizard Papillae Basilares

Iruanidae -

Central UHC2 increasing Basal

Sceloporus cyanogr'nys (Manley, '81)

increasing

frequency

Sceloporus orcutti (Turner et al., '81)

frequency CF' 4,300 Hz

CF 100-1,600 Hz CF groups 800, 1,800,2,500 Hz

Apical

Crotaphytus wislizeni (Manley, '81) 4,300 Hz 250-900 Hz

Anguidae Gerrhonotus multicarinatus (Weiss e t al.. '76. '78) , ,

Apical Basal CF 0.2-0.8 kHz 0.9 increasing frequency * 4.0 kHz

Tonotopic Teiidae

Tupinanbis nigropnnctc~tus (Browner and Caspary, '76) 0.4-1.3 kHz

Varanidae Varanus hengalensis (Manley, '77)

Apical Basal CF 1.3-2.8 kHz 0.2-1.0 kHz

Less sharply tuned Tonotopic, more sharply tuned Gekkonidae

Gekko gecko (Eatock and Manley, '81; Eatock et al., '81) Apical Middle Basal

Neural 3-5 kHz up to 2.5 kHz Low frequency Abneural 0.2 kHz

CF 0.1-4.0 kHz Frequency sensitive to a high degree Coleonyx nariegatus Cochlear Nucleus (Suga and Campbell, '67)

Scincidae Tmchysaurus rugosus (Johnstone and Johnstone, '69)

Mahuyo multicarinata (Manley, '81) CF 0.7-3 kHz

CF UD to 6.3 kHz

'Characteristic frequency 'Unidirectional hair cells.

their auditory papillae are relatively primitive in structure. such as those mentioned in 5 and 6 above could have been The papillae basilares of Sphenodon and turtles (Wever, derived from a basic teiid type of auditory papilla. '78; Miller, '78a) are moderately long (ca. 1 mm) and consist entirely of abneurally oriented hair cells that are covered ACKNOWLEDGMENTS completely by an unspecialized tectorial membrane.

(2) None of the lizards studied to date possesses a papilla basilaris composed entirely of abneurally oriented hair cells (Wever, '78; Miller, '80).

(3) However, I have studied and described three species of lizards with very large percentages (85-95%) of abneurally oriented hair cells, all of which are covered by an unspecialized tectorial membrane. These are species of teiid lizards, Ameiva ameiva, described before (Miller, '73b), and Cnemidophorus motaguae and Cnemidophorus tigris, described in this paper. (4) The hair cells of the above named teiid species are

probably innervated in a manner similar in many respects to that of the turtle (Beck and Miller, unpublished observations).

(5) The auditory papillae of the iguanid-agamid-anguid type, once thought to be primitive are probably specialized in that the bidirectional hair cells are well ordered and have lost all tectorial covering.

(6) Similarly, the auditory papillae of the scincid-gekkonid type have papillae with well ordered areas of bidirectionally oriented hair cells covered by very specialized types of tectorial membranes.

Therefore, I propose that the auditory papillae of certain species of the family Teiidae exhibit a structure more closely related to the probable ancestral stock than that of other extant lizards, and that other specialized types of papillae

The author is greatly indebted to Janet Beck for expert technical as well as collaborative contributions, to Dr. Jeanne Miller for participation in measuring and counting nerve fibers and for collation and organization of data, to David Akers and Simona Ikeda for expert photographic help, to The Coleman Laboratory of the Department of Otolaryngology for use of their light and transmission electron microscope facilities, to Dr. John Long for consultation in the interpretation of ultrastructure and for aid in the computerization of statistical programs, and to the depart- mental office staff. This work was supported by a USPHS grant No. RO 1 NS 11838.

LITERATURE CITED Arnesen, A.R., and K.K. Osen (1978) The cochlear nerve in the cat: Topog-

raphy, cochleotopy, and fiber spectrum. J. Comp. Neurol. 178:661-678. Bagger-Sjoback, D. (1976) The cellular organization and nervous supply of

the papilla basilaris in the lizard, Calotes uersicolor. Cell Tissue Res. 165141-156.

Baird, I.L. (1974) Anatomical features of the inner ear in submammalian vertebrates. In W.D. Keidel and W.D. Neff (eds): Handbook of Sensory Physiology, Vol. Vil. New York: Springer, pp. 159-212.

Bohne, B.A. (1976) Safe levels for noise exposure? Ann. Otol. 85:711-724. Bohne, B.A., A. Kenworthy, and C.D. Carr (1982) Density of myelinated

nerve fibers in the chinchilla cochlea. J. Acoust. SOC. Am. 72:102-107. Boord, R.L. (1969) The anatomy of the avian auditory system. Ann. N.Y.

Acad. Sci. 167:186-198.

24 M.R. MILLER

Boord, R.L., and G.L. Rasmussen (1958) Analysis of myelinated fibers of the acoustic nerve of the chinchilla. Anat. Rec. 130:394.

Bredberg, G. (1968) Cellular pattern and nerve supply of the human organ of Corti. Acta Otolaryngol. (Stockh.) [Suppl.] 236:l-135.

Browner, R., and D.Caspary (1976) The neurobiology of the acoustic tubercle in Tupinambis nigropunctatus. Soc. Neurosci. 6th Ann. Mtg. (Abstr.) 2:(1)15.

Corwin, J.T. (1981) Postembryonic production and aging of inner ear hair cells in sharks. J. Comp. Neurol. 201:541-553.

Corwin, J.T. (1983) Postembryonic growth of the macula neglecta auditory detector in the ray, Raja eleuata: Continual increases in hair cell number, neural convergence, and physiological sensitivity. J. Comp. Neurol. 217:345-356.

Dunn, R.F. (1978) Nerve fibers of the eighth nerve and their distribution to the sensory nerves of the inner ear in the bullfrog. J. Comp. Neurol. 182:621-636.

Diiring, M. von, A. Karduck, and H.-G. Richter (1974) The fine structure of the inner ear in Caiman crocodilus. Z. Anat. Entwickl. Gesch. 145:41- 65.

Eatock, R.A., G.A. Manley, and I,. Pawson (1981) Auditory nerve fiber activity in the Tokay Gecko: I, Implications for cochlear processing. J. Comp. Physiol. 142203-218.

Eatock, R.A., and G.A. Manley (1981) Auditory nerve fiber activity in the Tokay Gecko: 11, Temperature effect on tuning. J. Comp. Physiol. 142:219-226.

Ehret, G. (1979) Quantitative analysis of nerve fiber densities in the cochlea of the house mouse (Mus musculus). J. Comp. Neurol. 183:73-88.

Ehret, G., and M. Frankenreiter (1977) Quantitative analysis of cochlear structures in the house mouse in relation to mechanisms of acoustical information processing. J. Comp. Physiol. 122:65-85.

Engstrom, H., and B. Rexed (1940) Uber die Kaliherverhaltnisse der Ner- vendfasern im N. Statoacusticus des Menschen. Z. Mikrosk. Anat. Forsch. 47:448-455.

Etheridge, R. (1964) The skeletal morphology and systematic relationships of Sceloporine lizards. Copeia 610-631.

Fay, R.R., and A.N. Popper (1980) Structure and function in teleost auditory systems. In A.N. Popper and R.R. Fay (eds): Comparative studies of Hearing in Vertebrates. Berlin: Springer. pp. 3-42.

Gacek, R.R., and G.L. Rasmussen (1961) Fiber analysis of the statoacoustic nerve of guinea pig, cat, and monkey. Anat. Rec. I39:455-463.

Goodley, L.B., and R.L. Boord (1966) Quantitative analysis of the auditory papilla of the pigeon. Am. Zool. 6:542.

Hardy, M. (1938) The length of the organ of Corti in man. Am. J. Anat. 62:291-311.

Holton, T., and A.J. Hudspeth (1983) A micromechanical contribution to cochlear tuning and tonotopic organization. Science 222:508-510.

Johnstone, J.R., and B.M. Johnstone (1969) Unit responses from the lizard auditory nerve. Exp. Neurol. 24:528-539.

Leake, P.A. (1977) SEM observations of the cochlear duct in Caiman croce dilus. In: Scanning Electron Microscopy, Roc. Workshop Biomed. A p p L SEM Studies of Sensory Organs. Chicago: IIT Research Inst., Vol. 11, pp. 437-444.

Lewis, E.R., and Li, C.W. (1973) Evidence concerning the morphogenesis of saccular receptors in the bullfrog (Rana catesbeiana), J. Morphol. 139:351-362.

Lewis, E.R., E.L. Leverenz, and W. Bialek (1985) The Vertebrate Inner Ear. Boca Raton: CRC Press (in press).

Li, C.W., and E.R. Lewis (1974) Morphogenesis of auditory receptor epithelia in the bullfrog. In: Scanning Electron Microscopy, Part 111. Chicago: IIT Res. Inst., pp 791-798.

Manley, G.A. (1977) Response patterns and peripheral origin of auditory nerve fibers in the monitor lizard, Varanus bengalensis. J. Comp. Phys- iol. 118:(A)249-260.

Manley, G.A. (1981) A review of the auditory physiology of reptiles. In H. Autrum, D. Ottoson, E. Perl, and R.F. Schmidt (eds): Progress in Sensory Physiology. New York: Springer, pp. 49-134.

Miller, M.R. (1966) The cochlear duct of lizards. Proc.Calif. Acad. Sci. 33255- 359.

Miller, M.R. (1937a) A scanning electron microscope study of the papillar basilaris of Gekko gecko. Z. Zellforsch. 136:307-328.

Miller, M.R. (1973a) A scanning electron microscope study of the papillar basilaris of Gekko gecko. Z. Zellforsch. 136:307-328.

Miller, M.R. (1973b) Scanning electron microscope studies of some lizard basilar papillae. Am. J. Anat. 138t301-330.

Miller, M.R. (1974) Scanning electron microscope studies of some skink papillae basilares. Cell Tissue Res. 150:125-141.

Miller, M.R. (1978a) Scanning electron microscope studies of the papilla basilaris of some turtles and snakes. Am. J. Anat. 151:409-435.

Miller, M.R. (1978b) Further scanning electron microscope studies of lizard auditory papillae. J. Morphol. 156:381-418.

Miller, M.R. (1980) The reptilian cochlear duct. In A.N. Popper and R.R. Fay (eds): Comparative Studies of Hearing in Vertebrates. Berlin: Sprin- ger, pp. 169-204.

Miller, M.R. (1981) Scanning electron microscope studies of the auditory papillae of some iguanid lizards. Am. J. Anat. 162:55-72.

Morrison, D., R.A. Schindler, and J. Wersall (1975) A quantitative analysis of the afferent innervation of the organ of Corti in the guinea pig. Acta Otolaryngol. (Stockh.) 79: 11-23.

Ramprashad, F. (1976) Population and density of the bipolar ganglion cells in the cochlea of the harp seal (Pagophilus groenlandicus, Erxleben). Can. J. Zool. 54:1918-1926.

Ramprashad, F., K.E. Money, J.P. Landolt, and J. Laufer (1978) A neuroan- atomical study of the cochlea of the little brown bat (M.yotis lucrfugus). J. Comp. Neurol. 178:347-364.

Rasmussen, A.T. (1940) Studies ofthe VIIIth cranial nerve of man. Laryn~ goscope 5037-83.

Retzius, G. (1884) Des Gehororgen der Wirbeltiere. I1 Das Gehororgan der Rrptilien, der Vdgel und der Saugetiere. Stockholm: Samson und Wallin.

Romer, AS. (1966) Vertebrate Paleontology. Chicago: University of Chicago Press.

Savage, J.M. (1958) The iguanid lizard genera Urosaurus and Uta with remarks on related groups. Zoologica 43.4-54.

Smith, C.A. (1981) Recent advances in structural correlates of auditory receptors. In D. Ottoson (ed): Progress in Sensory Physiology, 2nd ed. New York: Springer, pp. 135-187.

Sneary, M. (1984) The structure of the turtle auditory receptor: A light, SEM, and TEM study. Anat. Rec. 208:171A-I72A.

Suga, N., and H.W. Campbell (1967) Frequency sensitivity of single auditory neurons in the gecko Coleonyx uariegatus. Science 157:88-90.

Teresi, P.V. (1984) Hair cell innervation patterns in the papilla basilaris of the western fence lizard, Sceloporus occzdentalis. Anat. Rec. 208:181A.

Turner, R.G., A.A. Murasaki, and D.W. Nielsen (1981) Cilium length: Influ- ence on neural tonotopic organization. Science 213:1519-1521.

Weiss, T.F., M.J. Mulroy, R.G. Turner, and C.L. Pike (1976) Tuning of single fibers in the cochlear nerve of the alligator lizard Relation to receptor morphology. Brain Res. 115:71-90.

Weiss, T.F., W.T. Peake, A. Ling, and T. Holton (1978) Which structures determine frequency selectivity and tonotopic 0r;:anization of vertebrate cochlear nerve fibers? Evidence from the alligator lizard. In R.F. Naun- ton (ed): Evoked Electrical Activity in the Auditory Nervous System. New York: Academic Press, pp. 91-112.

Wever, E.G. (1978) The Reptile Ear. Princeton, N J Princeton University Press.

Wever, E.G., J. McCormick, J. Palin, and S.H. Ridgway (1971) The cochlea of the dolphin, Turszops truncatus. Proc. Natl. Acad. Sci. USA 682908- 2912.

Wever, E.G., J.G. McCormick, J. Palin, and S.H. Ridgway (1972) Cochlear structure in the dolphin, Lagenorhyncr~s obliyuidens. Proc. Natl. Acad. Sci. USA 69:657-661.

White, J.S. (1978) A light and scanning electron niicroscopic study of the basilar papilla in the salamander, Ambystoma tLgrmum. Am. J. Anat. I51:437-452.

Zug, G.R. (1971) The distribution and patterns of the major arteries of the iguanids and comments on the intergeneric relationships of iguanids. Smithsonian Contrib. Zool. 83r1-23.

quantitative studies of auditory hair cells and nerves in lizards

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