Aqueous humor Aqueous humor DynamicsDynamics

Dr Shalmali RautDr Shalmali Raut

Embryology

Eye:NeurectodermSurface ectodermMesoderm (surrounding optic vesicle)Neural Crest

Cross section of 11 weeks foetus – the angular region is poorly defined at this stage and is occupied by loosely arranged, spindle shaped cells. Schlemm’s canal is unrecognizable, and ciliary muscles and ciliary processes are not yet formed

• Peters anomaly: Normal Ant chamber• ?peripheral ant synechiae• ? Trabecular changes characteristic of aging• Congenital glaucoma• congenital glaucoma may be due to 1. Persistance of a cellular or acellular membrane

over the the angle 2. Failure of differentiation or alterations in the

differential growth rates. • Goniodysgenesis• failure of remodelling of the progenitor tissues of

the angle • presence of strands between iris & trabecular

mesh work or between iris & cornea

Anatomy• Ciliary body• Limbal region

• Limbus: transition zone- sclera, cornea• Inner surface:scleral sulcus, scleral spur• Trabecular meshwork: converts to- Schlemms

canal• TM inserts- ridge-Schwalbe’s line• Outflow:TM, schlemm’s canal, intrascleral

channels

• Ciliary body: attch to SS- potential space- supraciliary space/supra choroidal space

• CB: Triangular on cross section(rt angle) In c/s it is triangular in shape and anterior posterior

length is 4.6 to 5.2 mm nasally to 5.6 to 6.3mm temporally

The inner side of the triangle is divided into two parts

• 1. pars plicata (2 -2.5mm) --- corona ciliaris• 2. pars plana (5mm wide temporaly,3mm nasally)

--- orbicularis ciliaris• Ciliary processes-inner, ant of Pars Plicata

Triangular shape of the ciliary body. The muscle fibres appear red in contrast with the connective tissue. The scleral spur is clearly delineated from ciliary muscle in the region of trabecular meshwork

Iris inserts CB – variable structure visible between the root of the iris and scleral spur – Ciliary body bandLens- suspended by Zonules- separates vitreous, AqueousIris: separates into anterior chamber and post chamberAnterior chamber angle: formed by iris and cornea.

Gonioscopic appearance of angle

Biology of Aqueous humour Inflow

Functions of inflow and outflowIf Inflow= Outflow…..steady state

Complex interrelationship

Ciliary body organisation

• Ciliary body muscle• Ciliary body vessels• Ciliary processes• Autonomic innervation of ciliary body

At birth anterior posterior length of ciliary body are 2.6 to 3.5 mm nasally, 2.8 to 4.3 mm temporally and reach three fourth of the adult dimension by 24 months with a constant ratio of 25% pars plicata to 75% pars plana.

It is forward continuation of choroid at ora serrata In c/s it is triangular in shape and anterior posterior length is 4.6 to 5.2

mm nasally to 5.6 to 6.3mm temporally The outer side of the triangle lies against the sclera with a

suprachoroidal space in between.

Cellular organisation of the ciliary body and ciliary processes

Ciliary body muscle

Ciliary processes• Ciliary capillaries• Stroma• Ciliary epithelium

nonpigmented pigmented

• Capillaries occupy the center ,it has thin endothelium lining with false porous areas,basement membrane surrounds endothelium , mural cells and pericytes are located within basement membrane

• Thin stroma• Ground substances (MPS,protein ,solute of plasma,collagen type III)

• Pigment epithelium• Low cuboidal epithelial cells• Numerous melanin granules in the cytoplasma• Separated from stroma by atypical basement membrane

• Non-pigmented epithelium• Columnar cells• Separated from aqueous humor by basement membrane• Presence of aquaporin-1 channel • Metabolically active cells

Remnants of anterior vitreous

Scanning electron micrograph of the human ciliary processes

Minor processes

Major processes

Autonomic innervation of ciliary body

Sympathetic fibers synapse in the superior cervical ganglion and the postsynaptic fibers are distributed to the ciliary body vessels, no innervations of ciliary epithelium has been identified ,catecholamine released from the sympathetic nerve endings diffuse to the adrenergic receptors on the ciliary epithelium leading to increase in aqueous secretion

Parasympathetic fibers originates from Edinger Westphal nucleus to innervate the ciliary muscles .stimulation of these fibers leads to release of acetylcholine which stimulates cholinergic receptors on the ciliary muscle, which contracts leading

1 ) relaxation of zonules and change the shape of crystalline lens. 2 ) reduces resistance to conventional aqueous outflow as it opens trabecular meshwork

Mechanism of aqueous formation

Site of Aqueous humour production: ant portion of Pars Plicata along tips of crests of ciliary processes.

Changes seen:NPE:interdigitations, Rough E.R, Mitochondria –more

Capillary endoth:more fenestrations Thinner stroma

Aqueous humor:plasma within capillaries

Post chamber

Flows around lens

Through pupil

Anterior chamber

Temperature gradient-cooler towards the cornea

Convection pattern

Aqueous humor must transverse- capillary wall, Stroma, epithelial bilayer

Principal barrier: NPE cell membrane, junctional complexes

Transport Processes:mech Ultra filtration Diffusion Active transport

Osmotic flow

Ultra filtration

Process by which fluid and solutes can cross semi permeable membrane under a pressure gradient (capillary blood flow)-150ml of blood flows through the ciliary process each minutes- Twice the weight of the ciliary process- 4% of plasma filtered through

fenestration of capillary process

In Ciliary body, fluid movement is favored by hydrostatic pressure difference between capillary pressure and interstitial fluid pressure (IOP) and is RESISTED by the difference between the oncotic pressure of the plasma and aqueous humor.

Colloid concentration is high in ciliary process which favors movement of water from plasma into ciliary stroma but retards movement of water from stroma into posterior chamber.

Thus ULTRA FILTRATION helps fluid out of capillary into stroma of ciliary process.

Alone cannot account for volume of fluid into posterior chamber

Ultra filtration and active secretion works in tandem

Active transport Energy dependent process. Selective movement of substances against

electrochemical gradient across cell membrane. Majority (80% to 90%) of aqueous humor formation

depends on active secretion at the Non-pigmented ciliary epithelium.

Process Pressure dependent?

Energy dependent

Substances transported

Active secretion no yes Na (water follows), Cl, K, HCO3,

ascorbate, amino acids

Ultrafiltration yes no Water, small water soluble molecules

Diffusion no no Lipid soluble molecules

Diffusion

Movement of lipid soluble substances across the concentration gradient

Sufficient diffusion exchange occur with surrounding tissues.

Anterior chamber aqueous resembles plasma more closely than posterior chamber fluid

Blood-aqueous barrier

It is formed by zonula occludentes & zonula adherens located between inner non pigment epithelium of ciliary body & tight junctions of iris & capillary endothelial cells.

Composition

• Difference in composition across ant & post chambers:

• Active transport in Post chamber• Passive transport in Ant chamber

Factors causing breakdown of blood aqueous barrier

Trauma

mechanical injury of lens or iris contusion paracentesis

Endogenous mediators

histamine bradykinn prostaglandins and other eicosanoids serotonin acetylcholine

Neural activity

stimulation of V C.N.

Immunogenic activity

bovine serum albumin

Chemical injury

nitrogen mustard formaldehyde acid alkali

Miscellaneous bacterial endotoxins X radiation laser energy infrared radiation alfa melanocyte stimulating hormone

Character of aqueous humor

Physical character– Osmotic pressure: higher than blood– Specific gravity slightly more than water– PH : 7.53(>plasma)

– Ref. Ind. :1.36 – Anterior chamber aqueous : 0.25ml– Posterior chamber aqueous : 0.5ml– Rate of formation : 2.3 µl/min (about 1% of AC vol/min)

Biochemical Composition

Inorganic Ions Organic Anions Proteins Carbohydrates Glutathione & Urea O2 & Co2

components Mmol/kg watersodium 143.50(146)

potassium 4.55(4.8)

calcium 1.70

magnesium 0.78

chloride 109.50(111.8)

bicarbonate 33.60(24)

lactate 7.40(8.2)

pyruvate 0.66

ascorbate 0.96

glucose 6.93(8.3)

urea 7.00(7.3)

Aqueous Humor composition

Substance Mechanism of entry

sodium AT/UF/D

chloride AT/pH/conc.of Na.

potassium AT/D

Ascorbic acid AT

Amino acid AT

Bicarbonate AT/Linked to Na/diffusion into vitreous/decomposed to CO2

Glucose Low in aqueous/lost in vitreous/taken up by lens and cornea

Phosphate low

Pyruvate and lactate Increase due to glycolytic activity of avascular structure

Inorganic ions

Concentration of Na, K, Mg in Aqueous is similar to plasma.

Ca - Half that of Plasma. Excess of Chloride & deficit of Bicarbonate ions. Phosphate concentration is low.

Ascorbic acid - Secreted actively in aqueous. Important Antioxidant

Lactic acid - Most abundant Source - Glycolytic Metabolism of Intraocular tissues.

Hyaluronic acid - Absent in plasma. Source -From Vitreous (during normal process of GAG

removal).

Organic anions

Proteins - Almost Complete Absence of Protein in aqueous - Owing to BAB. (0.02G/100ML ) - Vast majority of aqueous protein is accounted for by Albumin & Globulin from Plasma with higher conc of Albumin. (A:G Ratio of Aqueous humor is higher) Immunoglobulin - Source-Local Production via iris Lymphocytes Plasma cells, IgG - 3MG/100ML. Crystallins Increased in eyes with Cataract

Carbohydrates Glucose in Aqueous humor is approx 80% of plasma Probably enters Aqueous by simple diffusion. Ionositol is 10times that of plasma.

Glutathione Glutathione - Important tripeptide Source-diffusion from blood by an active transport system in

ciliary epithelium.

Stabilizer of Redox state of aqueous.

Urea - 80% of plasma conc. - Effective in hyper osmotic infusion treatment for glaucoma.

O2 & Co2

- Source- blood supplied to ciliary body & iris. - Corneal endothelium is critically dependent upon aqueous O2 supply for active fluid transport that maintains corneal transparency. - Lens & endothelial lining of Trab. meshwork also derive their O2 supply from Aqueous humor.

Biology of Aqueous Humor Outflow

• Cellular organisation of ciliary pathway:• Scleral spur• Schwalbe’s line• Trabecular meshwork• Uveal meshwork• Corneoscleral meshwork• Juxtacanalicular tissues• Schlemms Canal• Intra-scleral channels• Episcleral, conjunctival veins

Scleral spur– Fibrous ring– Meriodinal section

appears as WEDGE– Anteriorly attached to

Trabecular meshwork– Posteriorly attached to

sclera,longitudinal portion of ciliary muscle

– CM- contracts-pulls SS posteriorly- increases width of intertrabecular space and prevent SC from collapsing

Schwalbe’s line

- Collagen and elastic tissues- Irregular elevation- 50-150 µm

Runs circumferentially arround the globe

Transition zone – Trabeculum to corneal

endothelium– Termination of DM– Insertion of TM into

corneal stroma

Trabecular meshwork– Triangular (sieve like)

shape– Apex at schwalbe’s line– Base at scleral spur– Extend from peripheral

cornea and DM to scleral spur posteriorly

– Flat interconnected perforated sheets

– Fused in 2-3 layers anteriorly and 12-20 layers posteriorly

Uveoscleral meshwork– Flat sheet with wide irregular

perforation(25 to 75 um)– Multiple plane

Corneo-scleral meshwork– 8-15 perforated sheets– 5-12µm thickness– Elliptical in shape– Not aligned – Continuous flow of aqueous is must– Collagen type I,III,IV,V,VI,VIII– Laminin, fibronectin, heparin etc…

Juxta- canalicular meshwork– Thin layer of tissue– 2-20µm thickness– Separates outer layers of Corneo-

scleral network from inner wall of Schlemm's

– GAG,GP,COLLAGEN I,III,IV,V,– Elastic fibers

Actions– Passage of aqueous– Phagocytosis by Trabecular endothelium– Structural support to Schlemm’s canal by SS,TM,CM

Schlemm’s canal– Endothelial lined, which

are irregular, spindle shaped and contains giant vacuoles.

– Runs circumferentially– Resembles lymphatic

channel– Single slit like tube– 190-370 µm

Fig. shows the wall of Schlemm’s canal (SC) and adjacent trabecular meshwork. The endothelial lining of the trabecular wall of Schlemm’s canal is very irregular; normally, the cells show luminal bulges corresponding to cell nuclei (N) and macro vascular configuration (MV)

Corneoscleral wall of SC is more compact than the trabecular wall

Pericanalicular connective tissue of variable thickness supports the endothelial lining of Trabecular wall

Collector channelSchlemm’s channel drains into collector channel (25-35) also termed intrascleral aqueous vessels

Complex system

DIRECT System

formed by 8 larger vessel which terminates directly into episcleral vein

INDIRECT System

formed by numerous fine collector channel which drains into Deep intrascleral plexus, mid intrascleral plexus and episcleral plexus – before finally draining into episcleral veins

Intra-scleral channels

Episcleral veins - most of the aqueous vessels drain into it. episcleral veins

via anterior ciliary and superior ophthalmic vein cavernous sinus

Cellular organisation of Uveal outflow pathway

• Uveoscleral outflow• Uveocortex outflow

Aqueous humor drainage

Conventional or canalicular (Schlemm's canal) Unconventional or uveoscleral.

Trans canalicular flow Uveal meshwork

Corneoscleral meshwork Juxtra cannalicular meshwork Endothelial linning of SC 20-30 collector channel

Intrascleral venous plexus episcleral plexus Epicsleral venous plexus Anterior ciliary plexus aqueous vein

Trans cannalicular

Sponge like One way valve Prevents entry of toxic

substance Phagocytosis like RE system Absence of clotting factors Fibrinolytic activity Self cleaning filter

Uveoscleral pathway

Iris, Ciliary muscle, sclera

Supraciliary, suprachoroidal space

5-25% of flow

Mechanism of aqueous transport across inner wall of Schlemm's canal

Vacuolation theory

Leaky pores in endothelial cells

Contractile microfilaments

Sonderman’s channels

Vesicles and larger vacuoles are seen in endothelium, Tripathi et al has suggested that these vacuoles open and close intermittently to transport aqueous from juxtacanalicular tissue to Schlemm's canal

Vacuolation theory

Low magnification electron micrograph of endothelial lining of Schlemm's canal (SC) shows that majority of vacuolar configuration (V) at this level have direct communication (in arrows) with sub endothelial extra cellular spaces containing aqueous humor

Electron micrography of a vacuolar structure that shows both basal and apical openings thus constituting vacuolar trans cellular channel

Measurement of aqueous humor

The rate of flow is 2-3µl/min Measured

– Pressure dependent methods» Tonography» Suction cup» perfusion

– Tracer methods» Photogamametry» Radiolabelled isotopes» Fluorescein» Fluroscinated dextrans

Outflow mechanics Poiseuille Hagen Formula

– which states that resistance to flow of blood in a vessel is inversely proportional to 4th power of radius, hence flow in that vessel is directly proportional to the 4th power of radius.

Ohm’s law (V=IR)- The flow of aqueous through the trabecular

meshwork(I) will depend on the pressure difference between the intraocular pressure and the EVP(V) divided by the resistance of the trabecular meshwork(R)

– Conductance is inverse of resistance=(I=VC) and TM conductance seems to decrease with increase IOP

Goldmann proposed rate of aqueous outflow (F) is proportional to the IOP(P0) – Episcleral venous pressure (Pv) --------------- F α (P0 - Pv)

Grant proposed F = C(P0 - Pv) C is coefficient of outflow facility

ESTIMATION OF FACTOR “ C “(micro litre/min/mm Hg) Tonography estimates c value by raising the IOP with the weight

of an indentation tonometer and observing the decay curve in IOP.The weight causes raised IOP,leading to increased outflow and change in aqueous volume which is inferred from FRIEDENWALD table relating volume change to Schiotz change

NORMAL “C” is 0.28 micro litre /min /mm Hg

Tonography Unit

Functions of aqueous humor

Maintains volume and IOP Nutrition for avascular ocular structure

• Posterior cornea• Trabecular meshwork• Lens• Anterior vitreous

Optical role Antioxidant Clearing Function

Factors affecting IOP

LONG TERM CHANGES

Heredity Age Gender Refractive error Ethnicity

Diurnal variation• Maximum pressure in morning hours due to

increase in cortisol level• In Sleep ,IOP half that of awakening hours due to

decrease level of circulating catecholamine

Postural variation Exertional influences Lid and Eye movement

Intraocular condition ( Anterior uveitis, Rhegmatogenous retinal detachment)

Systemic conditions -Arterial blood pressure -Systemic venous pressure -Hyperthermia -Hormone -Plasma osmolarity

Environmental conditions General anesthesia Alcohol Caffeine Tobacco Heroin and marijuana

Molecular mechanisms of outflow resistance in Trabecular meshwork

• GAG:Glycosaminoglycans• Glucocorticoids• Myocilin• Contractile myofilaments• Sulfhydrylgroups• Fibrinolytic activity• Pressure dependent changes• Other TM markers and function• Age related changes

Mechanisms of Drug actionDrug Aqueous

prod.Trabecular M

Uveoscleral

B-blockers ↓

Pilocarpine ↑

Adrenaline, Dipevefrine

↓? ↑ ↑

Brimonidine ↓ ↑?

Prostaglandins ↑? ↑

Carbonic anhydrase (-)

↓