J. Embryol. exp. Morph., Vol. 12, Part 4, pp. 673-691, December 1964Printed in Great Britain

Oogenesis in the guinea-pig

by J . M. IOANNOU1

From the Department of Anatomy, University of Birmingham

WITH TWO PLATES

INTRODUCTION

I T is by now thoroughly established that the process of oogenesis followsessentially the same pattern in all but a few of the mammalian species studied.Thus primordial germ cells first appear extra-gonadally and migrate—activelyor passively—to the genital ridges, where they give rise to oogonia (see Franchi,Mandl & Zuckerman, 1962). After undergoing an unknown number of mitoticdivisions, the germ cells enter the prophase of meiosis.

The sequence of cytological events associated with oogenesis and with theprophase of meiosis has been described on numerous occasions (e.g. Winiwarter1901, 1908, 1910, 1920a, b; Winiwarter & Sainmont, 1909; Kingery, 1917;Allen, 1923; Evans & Swezy, 1931; Beaumont & Mandl, 1962; Baker, 1963;Hughes, 1963). The only adequate quantitative studies of oogenesis in mammals,however, are those of Mandl & Zuckerman (1951a), Mandl & Shelton (1959)and Beaumont & Mandl (1962) on the rat; Jones (1957), Jones & Krohn (1961)and Peters & Levy (1964) on the mouse; and Baker (1963) on human ovaries. Ineach of these species it has been shown that the process of oogenesis (i.e. theformation of new germ cells by the mitotic activity of oogonia) ceases before orshortly after birth. When the germ cells have passed into the prophase of meiosis,they become, by definition, oocytes. With the passage of time, the stock ofoocytes is gradually depleted by atresia and ovulation, but never added to.

A number of authors have carried out qualitative studies on the ovary of theguinea-pig. No attempt has thus far been made, however, to count the germcells in this species. It has been claimed, on the basis of qualitative observations,that neo-formation of germ cells occurs in the guinea-pig after birth and evenafter the attainment of sexual maturity (Sun, 1923; Papanicolaou, 1924; Harman& Kirgis, 1938; Schmidt & Hoffman, 1941; Bookhout, 1945; Aron, Marescaux& Petrovic, 1952, 1954a, b). The present study was therefore designed (i) to

1 Author's address: Department of Anatomy, The Medical School, Birmingham 15, England.

674 J.M.IOANNOU

count the germ cells in the ovary of the guinea-pig from the time of sex differen-tiation to beyond the time of sexual maturity; (ii) to examine the sequence ofmorphological changes associated with oogenesis and meiotic prophase; and(iii) to correlate the qualitative and quantitative observations.

MATERIALS AND METHODS

Eighty-nine ovaries were examined. Forty-eight were derived from forty-fourembryos and foetuses aged 26, 28,29, 30, 35,40,41,45, 50, 52, 55, 60 and 65 dayspost coitum (p.c.) The remaining forty-one ovaries were removed from animalsaged 5, 10, 15, 20 and 35 days, and 2, 3, 6 and 12 months post partum (p.p.).

Adult females were housed with fertile males and samples of the vaginalcontents examined daily. The dates of fertile matings were recorded and usedto determine the ages of embryos and foetuses removed at autopsy.

The animals were killed by means of chloroform vapour, and ovaries removedand fixed as quickly as possible.

Histological procedures

Ovaries from specimens older than 35 days p.c. were dissected free from allsurrounding tissues prior to fixation. The whole of the caudal half of youngerembryos was fixed, and the gonads sexed on subsequent histological examination.The tissues were placed in Bouin's aqueous fluid for 2 to 24 hr., depending onthe size of the specimen. After fixation, the material was transferred to 70 percent, alcohol, dehydrated, and then embedded in paraffin wax. Serial sectionswere cut at 5 or 7 /x, and stained with Weigert's iron haematoxylin and 'chromo-trop 2R\

Quantitative procedures

Volumetric assessment of the numbers of germ cells

Estimation of ovarian volume. Every 10th section of each ovary was projectedon to paper at a magnification ranging from x 23 to x 180, depending on thesize of the ovary, and the outline of each section was drawn. The volume of theovary was then calculated by the planimetric method of Dornfeld, Slater& Scheffe (1942). In some cases the volumes of both ovaries of a pair wereestimated, and the mean values used in subsequent analyses.

Estimation of cell and nuclear volumes at different stages of development. Theoutlines of the germ cells and their nuclei at different stages of developmentwere drawn by means of a camera lucida at magnifications varying betweenx 2350 and x 2800. At some stages the nuclear membrane was either absent(dividing oogonia, both normal and atretic) or too indistinct (pyknotic oogonia;oocytes at leptotene and zygotene) to be drawn with confidence. Seventy suchdrawings were made for most stages and fifty for all others. On the assumption

Obgenesis in the guinea-pig 675

that both cell and nucleus are essentially spherical, the mean radius (r) was usedto calculate the volume (V) of the cell or nucleus by applying the formula

The numbers of germ cells in each age group

The method used was essentially similar to that described by Beaumont &Mandl (1962) and Baker (1963). Chalkley's (1943) technique was employed toestimate the proportion of germinal to non-germinal tissue, at least 1500 cellsin each ovary being classified as germinal or somatic. A differential count wasthen made, 1000 germ cells in each ovary being sub-divided into the followingcell types (seven normal and four atretic):

Normal. Oogonia in interphase, mitotic prophase and mitotic metaphase;oocytes in the leptotene, zygotene, pachytene and diplotene stages of meioticprophase.

Atretic. Pyknotic oogonia; oogonia degenerating in mitosis ('atretic divi-sions'); and oocytes degenerating in early meiotic prophase ( 'Z' cells; seeBeaumont & Mandl, 1962) and in the diplotene stage.

In order to calculate the population (N) of each of these types of germ cell, itwas first necessary to derive the value of the mean corrected cell volume (Mcv)for each specimen. This was obtained from the equation:

where C, and Cv represent the percentage and the volume respectively of eachcell type, and x is the total number of cell types observed in each ovary. Thevalue of Mcv was used in a second formula to derive n, the total number of germcells of all types:

V_ _Mcv

V denotes the volume of ovarian tissue occupied by germ cells, and is the productof total volume and percentage germinal tissue for each ovary.

Finally, N is given by:nxCt = N

In this expression, C,, the percentage of each germ cell type, was provided by thedifferential count.

The validity of the volumetric method of estimating the population of germcells was assessed by comparing calculated values for four animals with countsof germ cells made on every 20th or 40th section. In the latter estimate, thesections were examined under an oil-immersion objective, and every germ cellwhich was judged to be cut through or near its maximum diameter was counted.The totals thus arrived at were then simply multiplied by either 20 or 40 to obtainthe total population (Table 7). Despite the many sources of error inherent in

44

676 J. M. IOANNOU

both methods, the results are in fair agreement. As the numbers of germ cellsincrease, however, the difference widens. In view of the inaccuracies unavoidablein both methods, it is impossible to judge which of the two provides the morereliable estimate.

Counts of germ cellsThe numbers of normal and atretic oocytes in the ovaries of animals older

than 55 days p.c. were assessed by counts made on every 10th or 20th serialsection of the ovary (depending on the size of the ovary concerned), using thenucleolus as a 'marker' (see Mandl & Zuckerman, 1951&; Green & Zuckerman,1951, 1954). The total population of oocytes was then obtained by multiplyingthe counts by either 10 or 20.

RESULTS

Qualitative observationsNormal oogonia

Oogonia at interphase are distinguishable by their relatively large size andspherical nucleus (Plate 1, Fig. A). One or two large nucleoli are usually present,often with several smaller micro-nucleoli. The chromatin in the nucleus is diffuseand appears slightly granular in histological preparations.

At prophase of mitosis (Plate 1, Figs. B, E), the nuclear chromatin is organizedinto chromosomes, which frequently lie immediately below the nuclear mem-brane, thus leaving a relatively clear area in the centre of the nucleus. The nuclearmembrane disappears at metaphase, and no nucleoli are visible (Plate 1, Fig. C).

Normal oocytes

The peak population of oocytes at leptotene is found at 35 days p.c Duringthis stage, long, thin chromosomes appear out of the diffuse chromatin materialof the pre-meiotic oocyte (Plate 1, Fig. E). The chromosomes are beaded, andoften bound tightly together.

The predominant phase at 40 days p.c. is zygotene, when the chromosomes arethicker (Plate 1, Fig. F). Polarization may occur at this stage, giving rise to abouquet-like configuration. By 50 days p.c. the most common stage found ispachytene (Plate 2, Fig. G), when the chromosomes are thicker still, and eitherdistributed fairly evenly throughout the nucleus or—more often—polarized toone side.

Through each of these successive stages the germ cell has increased in size.But the greatest size by far is reached at the diplotene stage (Plate 2, Fig. I),which first appears at 35 days p.c. The chromosomes at this stage are long andthin. Their outline is blurred, and lateral projections may often be observed.Chiasmata are frequently seen. The large nucleoli reappear and the nuclearmembrane becomes more distinct.

By the 60th day of intra-uterine life, some oocytes at the diplotene stage are

J. Embryol. exp. Morph. Vol. 12, Part 4

10/1

EXPLANATION OF PLATES

Histological preparations (Figs. A & J 7/x; B to I 5JU)

PLATE 1

FIG. A. Oogonia at interphase (30 daysp.c).FIG. B. 'Atretic division' (A); oogonium at prophase of mitosis (O) (30 days p.c).FIG. C. Oogonium at metaphase of mitosis (30 days p.c).FIG. D. Oogonium at anaphase of mitosis (30 days/?.c).FIG. E. Oocytes at Jeptotene (L); oogonia at prophase of mitosis (O) (35 days p.c).FIG. F. Oocytes at zygotene (41 days p.c).

J. M. TOANNOU {Facing page 676)

/ . Embryol. exp. Morph. Vol. 12, Part 4

PLATE 2

FIG. G. Oocytes at pachytene (52 days/?.c).FIG. H. 'Z ' cells, at zygotene (Z) and at pachytene (P) (50 days p.c).FIG. I. Large oocytes at diplotene (55 days/J.c).FIG. J. 'Contracted' oocyte at diplotene in a primordial follicle (3 months p.p).J. M. 10ANN0U (Facing page 677)

Oogenesis in the guinea-pig 611

already invested in large follicles (four or more layers of granulosa cells); by65 days, the ovaries contain a few Graafian follicles (five or more layers ofgranulosa cells surrounding an antrum).

Oocytes at the diplotene stage are predominant up to about 20 days p.p.Thereafter, the number of'normal' oocytes at this stage progressively decreases.A 'new' type of oocyte, encountered only in very small numbers before puberty,gradually increases in incidence as the animal ages. This cell displays a tightlybound ball of chromosomes, somewhat reminiscent of the foetal leptotene stage,and about the same size, but made up of much thicker chromosomal threads(Plate 2, Fig. J). Most of these oocytes show none of the characteristics ofatresia—the chromosome outlines are sharp and clear, the nuclear membrane isusually smooth, the cytoplasm may be slightly granular in appearance but israrely eosinophilic. With occasional exceptions, therefore, these cells areclassified as normal. Intermediate stages of contraction, between the normallarge diplotene oocyte and the fully 'contracted' stage, are also seen in animalsolder than 10 days.

Degenerating oogoniaOogonia may degenerate either at interphase, when they become pyknotic,

or while undergoing mitotic division. In the latter case they are known as'atretic divisions'.

In pyknotic oogonia, the nucleus condenses into a densely staining clump ofchromatin, the nuclear membrane may become wrinkled, and the cytoplasmusually becomes eosinophilic. In the late stages of pyknosis the nucleus may benothing more than a dark spherical mass of chromatin.

Atresia at mitotic prophase is characterized in its early stages by a condensa-tion of the chromosomal material, frequently onto the nuclear membrane. Atmetaphase the chromosomes appear to thicken, and their outlines becomeblurred. Advanced stages of degeneration are characterized by condensation ofthe nuclear chromatin into one dense mass, often with a raspberry-shaped outline(Plate 1, Fig. B).

'Z ' cellsThe chromosomes in these cells become blurred in appearance, and often

aggregate into one mass (Plate 2, Fig. H). The cytoplasm becomes highlyeosinophilic, and acquires a thick, granular character.

Atretic oocytes in the diplotene phase

The chromosomes tend to condense and clump together, while the cytoplasmbecomes eosinophilic. When the oocyte is enclosed in a Graafian follicle, thegranulosa cells frequently show the first signs of atresia. The nuclei of cellslining the antrum tend to become pyknotic, and the cells then often flake off intothe antrum.

678 J. M. IOANNOU

Quantitative studiesOvarian volume

The volume of the ovary of the embryonic and foetal guinea-pig increasessteadily from 0-09 mm.3 at 26 days p.c. to 1-02 mm.3 at 55 days p.c. (Table 1).The anomalous values at 45 and 52 days p.c. are unlikely to be significant.

Age (daysp.c.)2628293035404145505255

No. ofspecimens

11155425715

TABLE 1

Mean ovarian volume

No. ofovaries

11195426715

Mean ovarianvolume (mm.3)

0 0 90 1 10 0 60-180-440-520-761-120-980-691-02

Range(mm.3)

———

0-13-0-230-18-0-630-35-0-820-60-0-920-83-1-520-73-1-21

—0-79-1-30

Sizes of germ cellsThe volume of the germ cells increases progressively to the diplotene stage

(Table 2). It is worth noting, however, that oocytes at the pachytene stage areapparently smaller than those at the preceding zygotene stage. Each cell stagewas drawn from ovaries at the age group at which it occurred most frequently,with the qualification that oocytes at diplotene were drawn from ovaries of thegroup killed at 55 days p.c. Diplotene oocytes in post-natal animals grow largerstill, but their inclusion in the volumetric calculations would have been mis-leading. The assumption was made, based on qualitative observations, thatnormal and atretic oocytes at the diplotene stage have essentially the same volume.

Numbers of germ cells

As assessed by Chalkley's (1943) procedure, the proportion of ovarian tissueoccupied by germ cells increases from 15-5 per cent, at 26 days p.c. to a peak of20 • 7 per cent, at 41 days/?.c.; thereafter it declines to a minimum of 10 • 3 per cent,by 52 days^.c. (Table 3). The higher value at 55 days p.c. may be explained bythe increased proportion of large oocytes at the diplotene stage.

The proportion of atretic germ cells increases steadily to a peak of 3 • 7 per cent,of the ovarian volume at 40 days p.c, and subsequently declines to 1 • 5 per cent,by 55 days/7.c.

The absolute volume of ovarian tissue occupied by normal and degeneratinggerm cells is shown in Table 4 and Text-fig. 1. The volume occupied by normal

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680 J. M. IOANNOU

germ cells increases to a peak at 41 daysp.c, and then declines until 52 daysp.c.;it subsequently rises again, presumably because of the increased incidence oflarge oocytes at diplotene. The peak volume for degenerating germ cells occursat 45

1000 -

30 35 40 45

Age (Days p.c.)

50 55

TEXT-FIG. 1. Mean ovarian volume and volume of ovarian tissue occupied by germ cells.o o, mean ovarian volume; • • , total volume of germinal tissue; x x, volumeoccupied by normal germ cells.

The mean percentages of germ cells at the different developmental stages inthe embryonic and foetal guinea-pig are shown in Text-fig. 2, and the corres-ponding absolute numbers in Table 5. Text-fig. 3 shows the incidence of degener-ating germ cells throughout embryonic and foetal life, and Table 6 the numbers of

Oogenesis in the guinea-pig 681

germ cells per ovary in post-natal guinea-pigs. It will be seen that the numberof germ cells per ovary reaches a peak of almost 105,000 at 41 days p.c, and

TABLE 3

Mean percentage cell-types in embryonic and foetal ovaries {assessed by Chalkley'sprocedure)

Age{days p.c.)

2628293035404145505255

No. ofovaries

11175425515

Mean no.of cells

'hit'

20312920221120442180224016052579291530692737

Somatic(%)

84-5489-4285-7983-6685-7284-6379-2787-6789-4189-7388-49

Normalgerm cells

{%)14-389-76

13-3115-1712-7011-7217-219-408-889-299-99

Atreticgerm cells

{%)1 0 80-820-911-171-593-653-522-931-710-981-51

Totalgerminaltissue (%)

15-4610-5814-2116-3414-2915-3720-7312-3310-5910-2711-50

declines steadily beyond that age until only ca. 13,000 remain at 12 months/?./?.(Text-fig. 4). Of the 105,000 at 41 days/?.c, some 94,000 are normal. The propor-tion of normal germ cells fluctuates around 90 per cent, until about 5 days/7./?.,

TABLE 4

Calculated mean volume of germinal tissue per ovary

Volume of tissue (/x3 x 106) occupied by

Age{days p.c.)

2628293035404145505255

No. ofovaries

11175425515

Normal germcells

12-9410-748-38

28-2062-9964-07

13604103-0991 1064-10

126-29

Atretic germcells

0-970-900-572-257-50

18-1721-6432-8019-626-76

1319

All germcells

13-9111-648-95

30-4570-4982-24

157-68135-89110-7270-86

139-48

when it begins to decline. The number of 'normal' oocytes at the diplotene stagedecreases slowly until about 20 days/?./?., and then with increased rapidity until12 months p.p.; at this time only some 800-900 of the germ cells are 'normal'

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Oogeniesis in the guinea OO

684 J. M. IOANNOU

60-

20-

60-

° 2 0 -jiaoa1 60-

o 20-

§

60-

20-

60-

20-

Oogonia

Leptotene

Zygotene

Pachytene

"-"I Diplotene

Age (Days p.c.)

TEXT-FIG. 2. Percentages of oogonia and oocytes per ovary in embryonic and foetal guinea-pigs.

oocytes at the diplotene stage (Table 6). Their place is taken by the adult 'con-tracted' oocytes, the proportion of which increases steadily from 0-1 per cent,at 5 days p.p. to 94 per cent, by 12 months p.p. (Table 6 and Text-fig. 5).

Oogenesis in the guinea-pig 685

DISCUSSION

The present results clearly indicate that the process of oogenesis in the guinea-pig follows essentially the same pattern as has been described for the rat(Beaumont & Mandl, 1962), man (Baker, 1963) and the chick (Hughes, 1963).

40 45

Age (Days p.c.)

50 55

TEXT-FIG. 3. Estimated mean numbers of degenerating germ cells at different stages ofdevelopment, x x , pyknotic oogonia; • • , 'atretic divisions'; O O, 'Z 'cells; • • , atretic cells at diplotene.

The orderly sequence of events observed, leading from the mitotic proliferationof oogonia to the mature oocytes at diplotene found in the Graafian follicles, iscommon to these species, and in it there is no place for the neo-formation ofgerm cells after birth.

After their arrival in the gonads, the germ cells (oogonia) proliferate by mitosis,maximal mitotic activity being observed at 28 days p.c, and diminishing to zeroby 52 days p.c. The majority of germ cells enter the prophase of meiosis between

686 J. M. IOANNOU

30 and 35 daysp.c. At 41 days p.c, 36 per cent, of the germ cells are at zygotene,and by 50 days 38 per cent, are at pachytene. Thereafter the proportion ofnormal large oocytes at the diplotene stage increases greatly, subsequently

100-

.OH

*—x

30 40 50 60

Age (Days p.c)

2 3 6

Months p.p.

12

TEXT-FIG. 4. Numbers of germ cells per ovary in guinea-pigs from the time of sex differentiationto 12 months p.p. x x , mean values; • • , values for individual animals.

levelling off at over 90 per cent, until about 1 month after birth, when it beginsto fall. This decline is so rapid that by 2 months p.p. fewer than 20 per cent.,and by 12 months only 6 per cent, of the remaining germ cells are large oocytes

i 4 0

I 30£J> 20

I 10I

5 10 15 20 12

Age (Days p.p.) (Months p.p.)

TEXT-FIG. 5. Numbers of 'contracted' and non-contracted oocytes at diplotene per ovary inpost-natal guinea-pigs. • • , total germ cells; o o, non-contracted cells; x x ,' contracted' cells.

at diplotene. This sharp fall coincides with an increase in the proportion ofadult 'contracted' oocytes, which reaches a level of 94 per cent, at 12 months.

While the numbers of noimal germ cells are fluctuating in this way, four over-lapping waves of degeneration are also in progress. Thus the number of oogoniaundergoing pyknosis increases to a peak of over 1200 at 41 days p.c, and then

Oogenesis in the guinea-pig 687

gradually decreases to zero by the 60th day of intra-uterine life. Between 35and 45 days p.c. the number of 'atretic divisions' fluctuates around a peak ofsome 1700 before declining to zero in the next 15 days. ' Z ' cells are found inincreasing numbers until 45 days p.c, when their numbers start falling. A feware still found as late as 10 days/?./?. Oocytes degenerating at the diplotene stageare first observed at 45 days p.c., and their incidence increases in pace with thatof normal oocytes at diplotene; subsequently their number diminishes as thepercentage of 'contracted' cells increases.

TABLE 7

Numbers of germ cells in four ovaries, assessed by both the volumetric methodand by counts on every 20th or 40th section

Blockno.

1A

32

63

106

Age(days p.c.)

26

30

40

55

Numbers of germ cells

1. Volumetricmethod

12,545

19,899

49,376

37,339

2. Counts

15,400(1:20)

21,840(1:20)

55,880(1:20)

60,640(1:40)

1. Expressedas percentage of 2

81-46

91-11

88-36

61-57

The overlap observed in the development of the ovary of the guinea-pig isthus similar to that found in man (Baker, 1963) and in the chick (Hughes, 1963).A similar tendency has been reported for the rabbit (Winiwarter, 1901) and themouse (Borum, 1961). The close synchronization of development observedin the rat (Beaumont & Mandl, 1962) would thus appear to be an exceptionrather than the rule.

Beaumont & Mandl (1962) reported that the oocyte of the rat passes into theso-called dictyate stage shortly after entering diplotene. In this stage the chromo-somes become longer and finer, less distinct, and appear to form a reticular net-work. In man, on the other hand, the mature oocyte retains the chromatinconfiguration of the diplotene phase until the onset of the maturation division,without entering a dictyate stage. At this time lateral projections may be visible,apparently projecting from some of the chromomeres (Baker, 1963). In theguinea-pig, similarly, the chromosomes are clearly visible as definite threadsuntil contraction takes place. Sometimes, however, the outline of the threadsis 'furry', and lateral projections can be observed. These chromosomes thusresemble the lampbrush chromosomes described in newts by Gall (1954),except that the lateral projections on the chromosomes of the guinea-pig cannever be resolved into loops. Once contraction has taken place these projectionsare no longer visible.

688 J. M. IOANNOU

Between the ages of 55 days p.c. and 35 days p.p. the normal large oocyte atdiplotene is predominant. One month later—and no new germ cells are formed inthe interval—cells in the contracted stage gain preponderance. They musttherefore be derived from the normal large oocytes at diplotene. This hypothesisis fully supported by studies of ovaries aged between 10 and about 60 days p.p.,which reveal the existence not only of increasing absolute and relative numbersof the fully contracted oocytes, but also of numerous germ cells showing varyingdegrees of nuclear contraction. Beyond this time, most of the surviving oocytesare fully contracted, and intermediate stages are accordingly much less common.

The 'contracted' oocyte of the adult guinea-pig has frequently been observedand described in the literature. Usually, however, it has been classified as anearly stage of meiotic prophase (e.g. see Clark, 1923; Evans & Swezy, 1931;Aron et al, 1952,1954a, b; Zuckerman, 1956). Hitherto, only Oakberg & Clark(1964) have suggested that it represents a modified form of the diplotene stage,a conclusion fully corroborated by the present study.

The only other mammalian species in which contraction has been reported tooccur is the rat (Mandl & Beaumont, 1964); slight contraction occurs after theoocyte enters the dictyate stage, and from the age of about 3 weeks onwards.As long ago as 1928, however, Kawaguchi reported that nuclear contractionoccurs in spermatocytes of the silkworm shortly after the onset of the diplotenephase. The chromatin aggregates in the centre of the nucleus, leaving a clear areaall around it; subsequently, it spreads out again into the more typical configura-tion of diplotene. This pattern of chromosomal changes is somewhat less clear-cut in the female.

It is an open question whether the germ cells which eventually mature andovulate are derived from the stock of' contracted' oocytes, or from the remainingnormal, non-contracted cells. At 12 months p.p. there are still over 1600 non-contracted oocytes at diplotene. The female guinea-pig normally sheds anaverage of four oocytes each 16-day cycle (Hermreck & Greenwald, 1964),which accounts for some 92 oocytes each year. A female guinea-pig may breedin a laboratory colony for up to 3 years (see Paterson, 1957). Some 1300 non-contracted oocytes at the diplotene stage thus remain, though a large proportionof these may be expected to undergo either contraction or atresia. It is equallypossible, however, that all the oocytes become contracted, and that some of themexpand again and gather more layers of follicular cells around them. Thesewould subsequently come under the influence of pituitary gonadotrophic hor-mones and either mature further and ovulate, or become atretic.

SUMMARY

1. Estimations were made of the numbers of germ cells in the guinea-pigovary from embryonic stages to maturity. A volumetric method was used toestimate the numbers of oogonia and oocytes in animals aged 26 to 55 days p.c,

Oogenesis in the guinea-pig 689

while counts were made of the numbers of germ cells present between 60 days p.c.and 12 months p.p.

2. Neither the mitotic activity of oogonia nor the successive stages of meioticprophase are closely synchronized.

3. The process of oogenesis ceases at about 50 days p.c. Large numbers ofoocytes are found in the leptotene stage at 35 days, in zygotene at about 40 days,and in pachytene at 50 days p.c. Thereafter the proportion of oocytes in thediplotene phase increases, so that by a few days after birth all the oocytes are inthis stage. By 1 month p.p., however, two types of oocyte in diplotene are foundin large numbers. The first is comparable to the corresponding stage in rat, manand the chick, while the second displays a very contracted nucleus. The lattertype rapidly gains predominance, and by 12 months p.p. 94 per cent, of the germcells belong to it.

4. The number of germ cells per ovary increases from about 27,000 at 30 daysp.c. to reach a peak of almost 105,000 by 41 days p.c.; subsequently it declines tosome 13,000 by 12 months p.p. This decline is caused by a steady process ofatresia which affects germ cells from the earliest stages of development.

RESUME

Uoogenese chez le cobaye

1. On a estime le nombre de cellules germinales dans l'ovaire du cobaye, apartir des stades embryonnaires jusqu'a la maturite. Une methode volumetriquea ete appliquee, pour evaluer le nombre d'oogonies et d'oocytes dans les animauxages de 26 a 55 jours p.c, tandis qu'on a compte le nombre de cellules germinalespresentes entre 60 jours p.c. et 12 mois p.p.

2. Ni l'activite mitotique des oogonies, ni les stades successifs de la prophasemeiotique, ne sont etroitement synchronises.

3. Le processus de l'oogenese prend fin vers le 50 erne jour p.c. On trouve ungrand nombre d'oocytes au stade leptotene a 35 jours, au zygotene a environ40 jours, et au pachytene a 50 jours p.c. Apres cela, la proportion d'oocytesau stade diplotene augmente, de sorte que quelques jours apres la naissance, tousles oocytes se trouvent a ce stade. Mais apres un mois p.p., on trouve en grandnombre deux types d'oocytes en phase diplotene. Le premier type est comparableau stade correspondant chez le rat, l'homme et la poule; tandis que le deuxiemefait preuve d'un noyau fort contracte. Ce dernier type prend rapidement ledessus, et apres 12 mois/?./?., 94% des cellules germinales lui appartiennent.

4. Le nombre de cellules germinales par ovaire augmente d'environ 27,000 a30 jours p.c. jusqu'au maximum de presque 105,000 a 41 jours p.c.; par la suite,il y a une diminution vers 13,000 a 12 mois p.p. Cette diminution est due a unprocessus continu d'atresie, qui affecte les cellules germinales des les stadesdu developpement les plus precoces.

690 J. M. IOANNOU

ACKNOWLEDGEMENTS

The expenses incurred in this study were defrayed from grants, made to Professor Sir SollyZuckerman, F.R.S., by the Population Council, Inc. and the Medical Research Council.

The author is grateful to Dr A. M. Mandl for her constant guidance and encouragement.

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(Manuscript received 21st May 1964)

45