Investigative Ophthalmology & Visual Science, Vol. 32, No ... · 1680 INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / April 1991 Vol. 32 Fig. 3. Oblique-tangential section of the posterior

Investigative Ophthalmology & Visual Science, Vol. 32, No. 5, April 1991Copyright © Association for Research in Vision and Ophthalmology

Posterior Attachment of Ciliary Muscle in Young,Accommodating Old, Presbyopic Monkeys

Ernsr Tomm, Elke Lurjen-Drecoll, Wilfried Jungkunz, and Johannes W. Rohen

The authors studied the posterior attachment of the ciliary muscle in seven young (3-10 yr) and five old(26-34 yr) rhesus monkeys by light microscopy and electron microscopy. Posterior attachment of themuscle bundles consisted of elastic tendons, exclusively. The elastic tendons were continuous with theelastic lamina of Bruch's membrane and were also connected by smaller elastic fibers to an elasticmeshwork that surrounds the pars plana vessels. In some areas, the tendons formed focal contacts withthe endothelial cells. The authors found that in old eyes, the tendons and the elastic fibers of theposterior ciliary body showed pronounced structural changes. The tendons appeared thickened, showedincreased amounts of associated microfibrils, and were surrounded by dense layers of thick collagenfibrils. An increased amount of collagen fibrils was also seen between the elastic layer of Bruch'smembrane and the pigmented epithelium. A mechanical link between those collagen fibrils and theelastic fibers is suggested by the presence of osmiophilic points of contact. The age-related increase inelastic fibrillar material could cause decreased compliance of the posterior insertion of ciliary muscleand could be an essential factor for presbyopia in rhesus monkeys. Invest Ophthalmol Vis Sci 32:1678-1692, 1991

The function of the primate ciliary muscle in ac-commodation depends on the inward movement ofthe anterior muscle during contraction.1 The move-ment of the muscle requires a remodeling of its innerstructure. Morphometric studies on meridional sec-tions of human2 and monkey eyes,3'4 after treatmentwith pilocarpine and atropine show that contractionof the ciliary muscle is associated with an increase inarea of the circular and reticular portion and a de-crease in area of the meridional portion. Additionally,the meridional portion shortens and the posterior endof the ciliary muscle moves anteriorly. To support theanterior movement of the muscle, a firm anterior at-tachment is necessary. This is supplied by broad ine-lastic collagenous tendons, which insert within thescleral spur, and via the trabecular meshwork, into

From the Department of Anatomy, University of Erlangcn-Niirnberg. Erlangen, FRG.

Supported by grants 124/2-4 and 81/18-4 from the DeutscheForschungsgemeinschaft and the by grants EY 02698 andRR00I67 from the USPHS National Institute of Health.

Presented in part at the Annual Meeting of the Association forResearch in Vision and Ophthalmology, Sarasota, Florida, May1990.

Submitted for publication: May 22, 1990; accepted October 24,1990.

Reprint requests: Ernst Tamm, Anatomisches Institut der Uni-versitat Erlangen-Niirnbcrg, Lchrstuhl II, KrankenhausstraBe 9,D-8520 Erlangen, FRG.

the peripheral cornea.56'7'8 In contrast, the posteriorattachment consists of elastic tendons.2'9101 u 2 Due totheir extensibility, the contracting muscle can moveforward during accommodation. The stretched poste-rior elastic tendons pull the ciliary muscle backwardsduring relaxation and desaccommodation.

We have shown that with increasing age, the ciliarymuscle of the rhesus monkey (Macaca mulata) losesits anterior inward movement in response to pilocar-pine.4 Video recording of ciliary body coronal width,after central electric stimulation of young (3-yr-old)and old (>20 yr) iridectomized rhesus monkeys,shows loss of measurable ciliary body movement withage, indicating loss of effective ciliary muscle contrac-tion.13 The age-related decline in structural responseparallels the loss of functional accommodative re-sponse to pilocarpine or carbachol.l4 Thus, loss of cili-ary muscle movement seems to be involved in the lossof accommodation and the development of presbyo-pia in this species.

Age-related changes in the ultrastructure of the cili-ary muscle cells and intramuscular connective tissueof the rhesus monkey are moderate and do not ac-count for the loss of structural and functional re-sponses.15 An increased inelastic posterior fixation,however, might restrict ciliary muscle anterior inwardmovement. We, therefore, investigated the ultrastruc-ture of the posterior insertion of the rhesus monkeyboth in young (3-10 yr) and in old (26-34 yr) ani-mals.

167ft

No. 5 ATTACHMENT OF CILIARY MUSCLE IN PRESBYOPIC MONKEYS / Tomm er ol 1679

Fig. 1. Schematic drawing of the rhesus monkey ciliary body toshow the planes of orientation of the sections. Meridional, frontal(A), oblique-frontal (B), and oblique-tangential (C) sections werecut.

Material and Methods

The eyes of seven young adult (3-10 yr) and five old(26, 26, 30, 31, 34 yr) rhesus monkeys of both sexeswere studied. The monkeys were from caged coloniesof the Wisconsin Regional Primate Research Center(Madison, WI). The young animals were killed inconjunction with various nonocular protocols,whereas the elderly animals were debilitated and wereeuthanized. Anesthesia for in vivo enucleation andsystemic perfusion was intramuscular ketamine HC115 mg/kg and intravenous pentobarbital Na 25 mg/kg. The animals were killed by a pentobarbital over-dose. These experiments were conducted in accor-dance with the ARVO Resolution on the Use of Ani-mals in Research.

Three of the animals (8, 10, 34 yr) were perfusionfixed via the heart with Ito's fixative,16 after perfusionwith heparinized NaCl. The eyes of the remaining an-imals were fixed by immersion immediately after in

vivo enucleation. Windows were cut in the posteriorsclera and the cornea of these eyes, after which theentire eyes were placed in Ito's fixative for at least 3 hrbefore dissection, to preserve the architecture of theciliary muscle and its posterior attachment to Bruch'smembrane.4

The eyes were sent to us by Professor P. L. Kauf-man (Department of Ophthalmology, University ofWisconsin, Medical School, Madison). No ocular ab-normalities, other than senile cataract and peripheralcystoid retinal degeneration, were seen clinically orhistopathologically in the old animals. The old mon-keys received, in vivo, one drop of commercial oph-thalmic 10% pilocarpine hydrochloride topically inone eye and 1% atropine sulfate in the other eye, 45min and 30 min, respectively, before fixation began.In each case, the pupils responded appropriately. Inthe eyes of two of the old animals (26, 34 yr), we hadshown a marked decrease in or no morphologic re-sponse to pilocarpine.4

To ensure that the ciliary muscle remained an-chored anteriorly and via Bruch's membrane posteri-orly, four sectors running from the posterior to theanterior pole, with a width of approximately 5 mm atthe equator, were cut from each quadrant of the eyes.These specimens were embedded in paraffin in theusual manner. The rest of the globe was divided be-hind the ora serrata, and small pieces containing thewhole ciliary body, iris, adjacent cornea, and sclerawere cut. After postfixation in 1% osmium tetroxide,the specimens were dehydrated with graded alcoholsand embedded in Epon.

From both paraffin and Epon specimens, sectionsfor Iightmicroscopy were cut with different planes of

Fig. 2. Meridional section of the pos-terior attachment of the ciliary musclein a young rhesus monkey. At the poste-rior tip of a meridional muscle bundle(CM), an elastic tendon originates (ar-rowheads) that is continuous with theelastic lamina of Bruch's membrane(arrows). In the area of the posterior at-tachment, loose connective tissue withcapillaries (C), some fibroblasts, and me-lanocytes are present (semithin section,Richardson's stain, X53O).

1680 INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / April 1991 Vol. 32

Fig. 3. Oblique-tangential section of the posteriorattachment of the ciliary muscle in a young monkey.The elastic tendons (arrows) that originate from themuscle bundles (CM) are connected by smaller elas-tic fibrils to a network of elastic fibers (asterisk)surrounding the vessels of the posterior ciliary body.This network is continuous with the elastic layer ofBruch's membrane next to the pigmented epithelium(PE). Due to the obliquity of cut, the elastic lamina ispartly resolved into its substructure, which consists ofa meshwork of elastic fibers. In some areas, elastictendons are in close contact (arrowheads) with thewalls of the pars plana capillaries (C) (paraffin sec-tion, resorcin-fuchsin/van Giesson stain, XI000).

Fig. 4. Posterior attachment of themeridional ciliary muscle portion in rhe-sus monkey eyes. The meridional mus-cle bundles (CM) form elastic tendons(arrows) that are continuous with theelastic layer of Bruch's membrane. Ten-dons of different bundles are connectedby smaller elastic fibers to an elastic net-work that surrounds the vessels of thepars plana. Thick elastic tendons are inclose contact with the endothelium ofthe vessels.

No. 5 ATTACHMENT OF CILIARY MUSCLE IN PRESBYOPIC MONKEYS / Tamm er ol 1681

orientation, eg, meridional, frontal, oblique-frontal,and oblique-tangential (Fig. 1). Generally, the paraf-fin sections were 5 //m thick, and the Epon specimenswere 1 nm thick. In each plane, serial sections werecut through the whole muscle and the surroundingconnective tissue, both in paraffin and in Epon speci-mens. The paraffin sections were stained with a com-bined Weigert's resorcin-fuchsin/van Gieson stain

for elastic fibers and smooth muscle cells.17 Otherstandard elastic stains used were Gomori's aldehydefuchsin and VerhoefFs iron hamatoxylin.17 The Eponsections were stained with Richardson's stain18 andwith modified Weigert's resorcin-fuchsin and Ver-hoefFs iron hamatoxylin.19

Additionally, ultrathin sections were cut of the cili-ary muscle and its attachment to the surrounding

ET i

Fig. 5A. Oblique-tangential section of an elastic tendon (ET) of a ciliary muscle bundle (CM) form the meridional portion in a youngmonkey. In the region of the myotendinous junction, the muscle cells show tapered ends with long cytoplasmic processes and invaginations(arrows). The area of magnification is shown (circle) in Figure 5B (electromicrograph, X33OO),


connective tissue, using the planes of sectioning de-scribed above. The sections were treated with lead ci-trate and uranyl acetate. For electron microscopic in-vestigation, JEOL (JEM 100 b) and Zeiss (EM 902,Germany) electron microscopes were used.

Results

Young, Adult Animals

Lightmicroscopy: In the eyes of rhesus monkeys,the meridional portion of the ciliary muscle ends pos-teriorly about 0.5-0.9 mm anterior to the ora serrata.From the ends of the posterior-most muscle bundles,tendons originate that stain positive with all threeelastic fiber stains used. The band-like elastic tendonshave a smallest diameter of 0.6-0.8 nm, which is seenin meridional sections, and a widest diameter of 2-2.5nm. The tendons are continuous with the elastic lam-ina of Bruch's membrane (Fig. 2). The elastic tendonsof different bundles are connected by smaller elasticfibers (0.5-0.7 /zm) to an elastic network, seen in thestroma between ciliary muscle and pigmented epithe-lium. The elastic network surrounds the efferent ves-sels of the ciliary body in the pars plana and is con-nected with the elastic network of Bruch's membrane(Figs. 3, 4). In some areas, the elastic tendons are inclose contact with the pars plana capillaries (Figs. 3,4). In the posterior part of the reticular portion, themuscle bundles also form elastic tendons, which areconnected with an elastic network around the ciliarybody capillaries, and insert in the elastic lamina ofBruch's membrane. The elastic tendons are, however,shorter than those of the meridional portion.

Electronmicroscopy

Meridional Portion

In the region of the myotendinous junction, the in-dividual ciliary muscle cells show tapered ends withlong cytoplasmic processes and long furrows and in-vaginations (Figs. 5A, B). The elastic tendons origi-nate partly from the invaginations of the muscle cellsand partly from the outer surface of the tips (Fig. 5 A).Thus, the tips of the cells are completely embedded inelastic material. In regions, where the elastic tendonsare connected to the muscle cells, the cell membraneforms dense bands with adhering myofilaments. Inareas where the cell membrane is specialized by densebands, no intervening basal lamina is seen betweenthe elastic tendons and the cell membrane (Fig. 5B).

The elastic tendons show the typical ultrastructuralcharacteristic for elastic fibers. They consist of a cen-tral, amorphous, moderate osmiophilic core and acoat of microfibrils (Fig. 5).

This is the same for the fibers of the elastic network,which surround the capillaries of the ciliary body andare connected with the elastic tendons. The fibers ofthe elastic network are also connected with the basallamina of the endothelial cells by small microfibrils.Occasionally, the thick elastic tendons run close to thebasal lamina of the endothelial cells of the pars planacapillaries (Fig. 6A). In some areas, the elastic tendonscontact the endothelial cell without intervening basallamina. In these areas, the cytoplasm close to the cellmembrane of the endothelial cell is more electron-dense than elsewhere (Fig. 6B).

A small amount of loosely arranged collagen fibersembedded in an electron-lucent ground substance ap-

Fig. 58. Higher magnification of Fig-ure 5A. Where the elastic tendons (E)are connected to the muscle cells (C),the cell membrane forms dense bands(arrows), with adhering myofilaments.No intervening basal lamina is seen be-tween the elastic tendons and the cellmembrane (X6O,OOO).

No. 5 ATTACHMENT OF CILIARY MU5CLE IN PRE5DYOPIC MONKEYS / Tomm er ol 1683

Fig. 6A. Oblique-tangential section ofthe elastic tendons (ET) of the meridionalciliary muscle portion in a young monkey.The elastic tendons are in broad contactwith the basal lamina of the endothelialcells of the pars plana capillaries (arrows).The area of magnification (circle) isshown in Figure 6B (electronmicrograph,X3300).

Fig. 6B. Higher magnification of Figure6A, The elastic tendon forms close pointsof contact with the endothelial cells. Theelastic tendon appears to contact the cellmembrane directly, and the cytoplasmclose to the point of contact is moreelectron-dense than elsewhere (arrow,X60.000).

e

pears among the ends of the muscle bundles, the elas-tic tendons, the capillaries, and the elastic lamina ofBruch's membrane (Fig. 7). The collagen fibrils have adiameter of 40-50 nm. Near the muscle cells, the col-lagen fibers show no apparent association with theelastic tendons and pass randomly among the meshes

of the elastic network of the posterior ciliary body.Closer to Bruch's membrane, the collagen fibers areoccasionally in contact with small bundles of microfi-brils that emerge from protrusions of the elastic fibers.Between the elastic layer of Bruch's membrane andthe pigmented epithelium, micronbrils and small col-


Fig. 7. Oblique-tangentialsection of the posterior elastictendons (ET) in a young mon-key. The elastic tendons of dif-ferent bundles are connected bysmaller elastic fibers. Elastictendons show an intimate asso-ciation with the pars plana capil-laries (arrows). A small amountof loosely arranged collagen fi-

' brils (arrowheads) embedded inan electron-lucent ground sub-stance is seen (electronmicro-graph, XI 6,000).

lagen fibrils can be seen (Fig. 8). The microfibrils forma fine filamentous connection between the elasticlayer and the basal lamina of the pigmented epithe-lium (Fig. 8). Where the microfibrils are in contactwith the elastic layer, the elastic fibers form extensionsor protrusions (Fig. 8).

Reticular Portion

In the posterior part of the reticular portion, theelastic tendons are thinner, but they show the same

ultrastructural characteristics as in the meridionalportion. The myotendinous junction shows some dif-ferences to the meridional portion. The muscle cellsform a single invagination or furrow at their tip,which is filled with elastic material. The adjacent cellmembrane shows dense bands, whereas a basal lam-ina cannot be seen. The tendons are also seen adja-cent to endothelial cells of the capillaries of the ciliarybody and are connected to the basal lamina of theendothelial cells by microfibrils.

Fig. 8. Oblique-tangential section ofBruch's membrane and the pigmentedepithelium in the region of the attach-ment of the ciliary muscle tendons in ayoung monkey. Between the elasticlayer (EL) of Bruch's membrane andthe pigmented epithelium (PE), bun-dles of microfibrils and several colla-gen fibrils are seen. The microfibrilsemerge from protrusions of the elasticlayer (arrows) and extend towardthe basal lamina of the pigmentedepithelium, forming a fine filamen-tous connection (electronmicrograph,X 20,000).


Old Animals

Light Microscopy: In the three animals older than30 yr, neither the elastic tendons of the meridionalportion and the posterior reticular portion nor theelastic lamina of Bruch's membrane in the region ofits connection with the tendons can be stained withthe conventional histologic elastic fiber stains (Fig. 9).Also, when stained with Richardson's stain, the elasticfibers remain pale and do not stain intensely blue likethe elastic fibers in young animals (Fig. 10). This is,however, only true case for the region of the posteriorciliary body. In contrast, the elastic lamina of Bruch'smembrane posterior to the ora serrata and the elastic-like fibers of the trabecular meshwork show the samestaining pattern as in young animals. The elastic ten-dons in the old eyes are markedly thickened, with awidest diameter of 4.5-5.5 /im. The tendons are notsmooth as in young animals, but have a more irregu-lar shape, with fuzzy borders and a notched appear-ance (Fig. 9). The elastic fibers of the elastic networkin the posterior ciliary body are also thickened. Thenetwork does not show its normal architecture butappears disintegrated and fragmented (Fig. 9). Theconnective tissue between the elastic tendons of thereticular and meridional muscle bundles and the cili-ary pigmented epithelium is markedly thickened andhyalinized (Figs. 9, 10). This finding was especiallypronounced in the three animals older than 30 yr.

Electron Microscopy

Electron microscopic examination showed that allelastic fibers in the region of the posterior attachmentof the ciliary muscle are changed when comparedwith elastic fibers in the same region in young animals(Figs. 11-15).

The homogenous component of the elastic fibers iselectron lucent and is filled with fine fibrogranularmaterial. Additionally, it contains scattered bundlesof electron-opaque microfibrils, which coalesce todark, electron-dense areas (Fig. 11). The microfibrilshave a diameter of 10-12 nm and a periodicity of 15nm. In contrast to young animals, bundles of microfi-brils with similar diameter and periodicity are alsoseen at the attachment of the tendons to the musclecells. The microfibrils connect the homogenous com-ponent of the elastic tendon with the muscle cells (Fig.12A). Where dense bands are seen, they contact thecell membrane directly, without intervening basallamina (Fig. 12B). Near the muscular insertion, theelastic tendons are embedded in a dense, granular os-miophilic ground substance that contains scatteredcollagen fibers (Fig. 12A). This is not seen in younganimals. The amount of surrounding collagen in-creases posteriorly toward Bruch's membrane. In this

Fig. 9. Oblique-tangential section of the posterior attachment ofthe ciliary muscle in a 34-year-old monkey. Region and orientationof the section are comparable with Figure 3. The elastic tendons(arrows) that originate from the muscle bundles (CM) are thickenedand have an irregular shape, with fuzzy borders and a notched ap-pearance. Although a histologic stain for elastic fibers is used, nei-ther the elastic tendons nor the elastic lamina of Bruch's membrane(white arrows) stain positive. The connective tissue between theelastic tendons and the ciliary pigmented epithelium (PE) is thick-ened and hyalinized (asterisks) (semithin section, resorcin-fuchsinstain, XI000).

region, the elastic tendons are almost surrounded by acoat of collagen fibrils (Fig. 13). The collagen fibrilshave diameters of 200-250 nm (Fig. 13), are parallelto the elastic tendons, and pass through the meshes ofthe elastic network of the posterior ciliary body.Among the collagen fibrils, clusters of membrane-bound vesicles and granular and tubular structurescan be seen (Fig. 13). These connective tissue changesare all characteristic for old animals, but are most pro-nounced in the three animals older than 30 yr.

Between the elastic layer of Bruch's membrane andthe ciliary pigmented epithelium, there is a thick,dense collagen layer (Fig. I4A). The collagen fibrilsshow a mainly meridional orientation, runningroughly parallel to the sclera. The collagen fibrils areattached to the elastic lamina anteriorly and to the

1686 INVESTIGATIVE OPHTHALMOLOGY b VI5UAL SCIENCE / April 1991 Vol. 32

Fig. 10. Posterior attachment of ciliarymuscle in a 31-year-old rhesus monkey.The posterior ends of the meridionalmuscle bundles (CM) and the capillaries(C) are surrounded by a homogenous,hyalinized extracellular matrix. Theelastic lamina of Bruch's membrane(arrows) stains pale and is thickenedcompared with those of young animals.A marked hyalinization is seen (aster-isks) between elastic lamina and pig-mented epithelium, (semithin section,Richardson's stain, X53O).

thickened basal lamina of the pigmented epitheliumposteriorly. They are continuous with dense arrays ofmicrofibrils that emerge from the elastic lamina ofBruch's membrane (Fig. 14B). The microfibrils havethe same orientation as the collagen fibrils. In someareas, the collagen fibrils contact the elastic laminawithout intervening microfibrils. The collagen fibrilsappear to be anchored to the elastic layer by osmio-

philic material (Fig. 15). The alignment of the colla-gen fibrils suggests that they are under tension andexert a pull on the elastic lamina, allowing small pro-trusions. The collagen fibers extend toward the pig-mented epithelium and show an intimate associationwith its thickened basal lamina.

In the eyes of all of the old animals studied, therewas no difference in the ultrastructure of the posterior

f \

Fig. 11. Oblique-tangentialsection of a posterior elastic ten-don in a 34-year-old monkey.Compared with those of younganimals, this tendon showsmarked structural changes. Thehomogenous material of thetendon appears electron lucentand is filled with fine fibrogran-ular material (asterisk). Addi-tionally, it contains scatteredbundles of electron-opaque mi-crofibrils (arrows) that show aperiodicity and coalesce to darkelectron-dense areas (electron-micrograph, X44,000).

No. 5 ATTACHMENT OF CILIARY MU5CLE IN PRESDYOPtC MONKEYS / Tomm er dt 1687

Fig. 12A. Oblique-tangential sectionof the myotendinous junction of an elas-tic tendon in the region of the posteriorattachment of the meridional portion ofthe ciliary muscle in a 34-year-old mon-key. Muscle cell (C) and tendon aresurrounded by electron-dense, amor-phous, granular, extracellular material(asterisks) (electronmicrograph, X7500).

Fig. 12B. Higher magnifica-tion of Figure 10A. At the myo-tendinous junction, bundlesof microfilaments (asterisks)emerge from the homogenouscomponent of the elastic ten-don and extend deeply into theinvaginations formed by themuscle cell (C). Where the mi-crofilaments appear to be at-tached to the cell membrane,the basal lamina is interrupted(arrowheads) and dense bandswith adhering myofilaments(arrows) are formed (X40,000).

/

1688 INVESTIGATIVE OPHTHALMOLOGY G VISUAL SCIENCE / April 1991 Vol. 32

ET

Fig. 13. Oblique-tangential section ofa posterior elastic tendon in a 34-year-old monkey. The homogenous materialof the elastic tendon (ET) stains electronlucent and contains bundles of electron-opaque microfibrils (arrow). The tendonis almost surrounded by collagen fibrils(asterisks), that have a diameter of 200-250 nm. Between the collagen fibril clus-ters of membrane-bound vesicles, granu-lar and tubular structures can beseen (arrowheads) (electronmicrograph,XI 4,300).

attachment of the ciliary muscle. There was also nodifference in the ultrastructure of this region betweenpilocarpine-treated and atropine-treated eyes.

Discussion

This study shows that the posterior tendons of thereticular and meridional muscle bundles of the ciliarymuscle of the young rhesus monkey are purely elastic,with the typical histologic staining qualities and ultra-structural characteristics of elastic fibers found inother tissues.2021-22 Thus, the tendons differ ultrastruc-turally from the anterior elastic-like tendons of theprimate ciliary muscle,82324 that are seen in additionto the anterior collagenous tendons.56'7'8

Distinctive ultrastructural features of the posteriorciliary muscle tendons are the numerous projectionsand invaginations of the muscle cell membrane andthe elongated membrane-bound dense bands in themyotendinous region. These features are also typicalfor the collagenous tendons of striated musclecells.25'26'27 True elastic tendons of smooth musclecells, however, have been described for the humanarrector pili muscle2829 and the homologous feathermuscle of birds.30 In all of these elastic tendons, themuscle cells show similar ultrastructural features inthe region of the myotendinous junction, but themode of the elastic fiber attachment differs. In young

animals, the ciliary muscle, the homogenous part ofthe elastic fiber, is in close contact with the cell mem-brane, and a basal lamina cannot be seen. In contrast,in the skin and feather muscle tendons, dense arraysof the elastin-associated microfibrils are embedded inthe muscle cell basal lamina.2930

Elasticity of the posterior attachment of the ciliarymuscle is important to allow forward movement ofthe muscle in accommodation and subsequent resto-ration of its length and position during desaccommo-dation. Desaccommodation implies that the tensionis restored to the elastic components of the lens.Helmholtz31 has assumed that the elastic force thatcauses desaccommodation resides in the choroid,since desaccommodation is associated with relaxationof the muscle. This ultrastructural study shows howciliary muscle is linked via elastic tendons to tissuestructures that are responsible for the elasticity of thechoroid.

In the region of the posterior attachment of the cili-ary muscle, young animals show a connection be-tween the elastic lamina of Bruch's membrane andthe basal lamina of the pigmented epithelium. Similarmicrofibrillar connections between elastic materialand the basal lamina of the PE have also been re-ported for the posterior ciliary body in the bovineeye32 and in rabbits and humans.33 In the primate eye,this attachment might be responsible for the move-


Fig. 14A. Oblique-tangential sectionof Bruch's membrane and the pigmentedepithelium in region of the attachmentof the ciliary muscle tendons in a 34-year-old monkey. The elastic lamina ofBruch's membrane (asterisks) appearsthicker than those in young animals. Thehomogenous component of the elasticmaterial stains electron lucent. Betweenthe elastic lamina and the pigmented epi-thelium (PE), a high amount of collagenfibrils in an elelctron-dense ground sub-stance is seen. The collagen fibrils con-tact the basal lamina of the PE (arrows)and the elastic lamina. In areas of con-tact, the elastic lamina shows protru-sions (arrowheads). The area of magnifi-cation is shown (circle) in Figure 12B(electronmicrograph, X4000).

Fig. 14B. Higher magnification of Fig-ure I2A. Microfibrils (asterisk) emergefrom the elastic layer and seem to be incontinuity with thicker collagen fibrils,which show their typical periodicity(arrow) (X40,000).

B

ment of the retina and ora serrata during accommoda-tion in human34'35'36 and young monkey eyes37 andthe occasional retinal breaks and consecutive detach-ments after strong miotics.38'39'40'41

The elastic tendons of the ciliary muscle are con-nected to Bruch's membrane and, in some areas, tothe endothelium of the efferent vessels of the ciliarybody, where junction-like osmiophilic structures areseen. Such connections between the capillary and a

tendon have not been described. After prolongedtreatment of monkey eyes with phospholine, thesesame vessels were dilated.42

The elastic fibers in the region of the posterior at-tachment of the ciliary muscle undergo structural age-related changes. These changes are different fromthose, that have been described in the eye, eg, in theposterior parts of the choroid.43'44'45 We do not knowto what extend these changes influence the functional

1690 INVESTIGATIVE OPHTHALMOLOGY 6 VISUAL SCIENCE / April 1991 Vol. 32

Fig. IS. Oblique-tangential section ofthe elastic layer of Bruch's membrane inthe region of the attachment of the cili-ary muscle tendons in a 34-year-oldmonkey. The collagen fibrils, which runbetween pigmented epithelium and theelastic layer (E) in areas, contact the elas-tic fibers directly, without interveningmicrofibrils. At points of contact, osmio-philic material is seen between collagenfibrils and the protrusions of the elasticlayer (arrows) (electronmicrograph,X40,000).

properties of the fibers and whether they representelastic fibers in the true sense of the word. Furtherstudies are needed, eg, using antibody staining, to de-termine whether these fibers contain elastin.

A distinct feature other than the structural changesin the elastic fibers is the pronounced increase in theamount of microfibrils and collagen fibrils, which areclosely associated with the elastic fibers. A mechanicallink between elastic and those collagen fibrils is sug-gested by the presence of osmiophilic points of con-tact between elastic and collagen fibrils not seen inyoung animals. The contacts are not affected by acutetopical adminisstration of pilocarpine or atropine,which would affect the tension in the tissue. Thus, thefibrils have a mechanical strength. The reason for theage-related increase in fibrillar material in the regionof the posterior attachment of the ciliary muscle is notclear. It is possible that mechanical traction of theaccommodating muscle serves as a stimulus for theproduction of extracellular material that accumulatesduring the lifetime of the animal. It has been shown invitro that cyclic stretching stimulates synthesis of col-lagen and other matrix components by arterialsmooth muscle cells.46 Hypertrophy of intestinalsmooth muscle in vivo is associated with an increasein the content of collagen.47-48 Increase in inelastic col-lagenous material and its association with the elasticsystem might stiffen the posterior insertion of the cili-ary muscle with increasing age and contribute to pres-byopia. Interestingly, it has been suggested that athree-dimensional interlocking of both collagen and

elastic fibers may be the cause of decreased tissue com-pliance in aged skin.49

Age changes in individual ciliary muscle cells areminimal.15 However, the fact that the area of rhesusmonkey ciliary muscle in meridional sections de-creases by approximately one-third with age,4 mightindicate loss of contractile muscle cells. Thus, themuscle might not be able to overcome the increasedresistance of the posterior attachment. The contrac-tions of the muscle might become more isometric asthe muscle is restrained in a relaxed position.

Reports show, using ingenious but indirect meth-ods, that human ciliary muscle does not weaken withonset of presbyopia.50'51 However, forces on the lensduring accommodation are determined less by thecontractile strength of the ciliary muscle than by itsposition and configuration. Therefore, changes in elas-ticity of the posterior attachment of the ciliary musclemight also be an important factor that accounts forage-dependent loss of accommodation in humans.However, since differences have been reported in age-related morphologic changes of the ciliary body be-tween humans5253 and rhesus monkeys,'s further stud-ies are needed to determine clarify whether similarmechanisms affect human presbyopia.

In summary, we hypothesize that an age-related in-crease in the amount of inelastic fibrillar materialcauses decreased compliance of the posterior inser-tion of the ciliary muscle in rhesus monkeys. As aresult, the anterior-inward movement of the ciliarymuscle during accommodation diminishes with age.


Loss of ciliary muscle movement might be responsi-ble for or at least contribute to presbyopia in primates.

Key words: ciliary smooth muscle, presbyopia, elastic fibers,aging, rhesus monkey

Acknowledgments

This article is dedicated to Professor Ernst H. Barany onthe occasion of his 80th birthday.

The authors thank Elke Kretschmar and Gcrtrud Linkfor technical assistance, Marco GoBwein for preparation ofthe photographs, and for preparation of the drawings weremade by Christiane Wittek.

References1. Rohcn J W: Scanning electron microscopic studies of the zonu-

lar apparatus in human and monkey eyes. Invest OphthalmolVis Sci 18:133, 1979.

2. Rohcn JW: Ciliarkorper (Corpus ciliarc). In Handbuch dermikroskopischen Anatomic des Mcnschcn. Vol. 3. pi. 4, Haulund Sinncsorganc. Das Augc und seine Hilfsorganc.Mdllendorf Wv and Bargmann W, editors. New York.Springer-Verlag. 1964, pp. 189-237.

3. Liitjcn E: Histomctrische Untersuchungen iibcr den Ciliar-muskcl des Primaten. Gracfes Arch Klin Exp Ophthalmol171:121. 1966.

4. Liitjen-Drecoll C, Tamm E, and Kaufman P: Agc-relatcd lossof morphologic response to pilocarpine in rhesus monkey cili-ary muscle. Arch Ophthalmol 106:1591, 1988.

5. Rohcn J W: Ubcr den Ansatz der Ciliarmuskulatur im Bercichdes Kammerwinkels. Ophthalmologica 131:51. 1956.

6. Kupfer C: Relationship of ciliary body meridional muscle andcorneoscleral trabecular meshwork. Arch Ophthalmol 68:818.1962.

7. Rohcn JW, Liitjen E. and Barany EH: The relation between theciliary muscle and the trabecular meshwork and its importancefor the effect of miotics on aqueous outflow resistance. A studyin two contrasting monkey species. Macaco irus and Ccrco-piifiecus aclhiops. Graefes Arch Klin Exp Ophthalmol 172:23,1967.

8. Rohen JW and Liitjen-Drecoll E: Morphology of aqueous out-flow pathways in normal and glaucomatous eyes. In The glau-comas. Vol. 1, Ritch R. Shields MB. and Krupin T, editors. St.Louis. C. V. Mosby Co., 1989. pp. 41-74.

9. IwanofF A and Arnold J: Mikroscopische Anatomic des Uveal-traktus und der Linse. /// Handbuch dergesamten Augenheil-kunde. Vol. 1, Grade A and Sacmisch T, editors. Leipzig. Ver-lag von Wilhelm Engelmann. 1874. pp. 265-320.

10. Salzmann M: Anatomic und Histologiedcs menschlichen Au-gapfels im Normalzustande, seine Entwicklung und sein Al-lern. Leipzig, Wien, Franz Dcutickc. 1912. pp. 120-124.

11. Laubcr H: Der Strahlenkorpcr (Corpus ciliarc). In Handbuchder mikroskopischen Anatomic des Menschen, Haul und Sin-ncsorgane, Augc. Vol. 3. pt. 2. Mollendorf Wv, editor. Berlin,Springer-Verlag. 1936, pp. 134-176.

12. Rohcn J W: Der Ziliarkorper als funktionclles System. Gegen-baurs Morphol Jahrb 92:415, 1952.

13. Neidcr M. Crawford K. Kaufman PL, and Bito LZ: In vivovidcography of the rhesus monkey accommodative apparatus.Age-related loss of ciliary muscle response to central stimula-tion. Arch Ophthalmol 108:69, 1990.

14. Bito LZ, DeRousseau CJ. Kaufman P. and Bito JW: Age-de-

pendent loss of accommodative amplitude in rhesus monkeys:an animal model for presbyopia. Invest Ophthalmol Vis Sci23:23, 1982.

15. Liitjen-Drecoll E, Tamm E and Kaufman P: Age changes inrhesus monkey ciliary muscle: Light and electron microscopy.Exp Eye Res 47:885, 1988.

16. Ito S and Karnovsky MJ: Formalaldchyde-glutaraldehydc fix-atives containing trinitro compounds. J Cell Biol 39:168a,1968.

17. Romcis B: Mikroskopische Tcchnik. 17th ed. Bock P, editor.Munich, Baltimore, Urban & Schwarzenbeck, 1989, pp 517-522.

18. Richardson KC, Jarrct L, and Finkc H: Embedding in cpoxyresins for ultrathin sectioning in electron microscopy. StainTechnol 39:229. 1960.

19. Bock P: Der Scmidiinnschnitt. Miinchcn, FRG, BergmannVerlag. 1984, pp. 132-135.

20. Greenlcc TK, Ross R. and Hartman JL: The fine structure ofclastic fibers. J Cell Biol 30:59. 1966.

21. Ross R: The elastic fiber. A review. J Histochcm Cytochem21:199, 1973.

22. Franzblau C and Faris B: Elastin: In Cell biology of extracellu-lar matrix, Hay ED, editor. Plenum Press, New York, 1981, pp.65-89.

23. Liitjen-Drecoll E, Futa R, and Rohcn JW: Ultrahistochcmicalstudies on tangential sections of the trabecular meshwork innormal and glaucomatous eyes. Invest Ophthalmol Vis Sci21:563, 1981.

24. Liitjen-Drecoll F.. Shimizu T, Rohrbach M, and Rohcn JW:Quantitative analysis of "Plaque Material" between ciliarymuscle tips in normal- and glaucomatous eyes. Exp Eye Res42:457. 1986.

25. Hanak H and Bock P: Die Fcinstruktur der Muskcl-Sehncn-vcrbindung von Skelett-und Hcrzmuskel. J Ultrastruct MolStruct Res 36:68, 1971.

26. Korneliusscn H: Ullrastructurc of the myotendinus junctionsin myxine and rat. Z Anat Entwickl Gcsch 142:91, 1973.

27. Nakao T: Some observations on the fine structure of the myo-tendinous junction in myotomal muscles of the tadpole tail.Cell Tiss Res 166:241, 1976.

28. Wicnkcr HG: Elcktroncnmikroskopischc Untersuchungeniiber den M. arrccior pili des Menschen. Z Mikrosk AnatForsch76:l, 1969.

29. Rodrigo FG, Cotta-Pcrcira C and David-Ferrcira JF: The finestructure of the elastic tendons in the human arrectorpili mus-cle. Br J Dermatol 93:631. 1975.

30. Drenckhahn D and Jcikowski: The myotendinous junction ofthe smooth feather muscles (mm. pennati). Cell Tiss Res194:151, 1978.

31. Helmholtz H. Handbuch der Physiologischcn Optik. In Allge-meine Enzyklopedie der Physik, Vol. IX. Karstcn G, editor.Leipzig. VossL. 1856.

32. Strccten BW and Licari BA: The zonulcs and the clastic micro-fibrillar system in the ciliary body. Invest Ophthalmol Vis Sci24:667. 1983.

33. Korte GE and D'Aversa G: The elastic tissue of Bruch's mem-brane. Connections to the choroidal elastic tissue and the cili-ary epithelium of the rabbit and human eyes. Arch Ophthalmol107:1654. 1989.

34. Hollins M: Docs the central human retina stretch during ac-commodation? Nature 251:729, 1974.

35. Enoch IM. Retinal stretch and accommodation. In Currentconcepts in ophthalmology. Vol. 5. Kaufman HE and Zim-merman TJ, editors. St. Louis, C.V. Mosby Co.. 1976, pp. 59-78.


36. Moses RA. Accommodation. In Adler's physiology of the eye:Clinical Application, Moses RA and Hart WM Jr, editors. St.Louis, C.V. Mosby Co., 1987. pp. 291-310.

37. Maurcr J: Histometrische Untersuchungen iiber Lage-vcranderungen der Ora scrrata bei vcrschiedenen Kontrak-tionszustanden des Ziliarmuskels im Primatenauge. Inau-gural-Dissertation der Medizinischen Fakultat dcr Friedrich-Alexander-Univcrsitat Erlangen-Niirnberg, Erlangen, FRG,1984.

38. Marr WC: The clinical use of di-isopropyl fluorophosphate(D.F.P.) in chronic glaucoma. Am J Ophthalmol 30:1473.1947.

39. Leopold 1H and Me Donald PR: Di-isopropyl fluorophosphate(DFP) in treatment of glaucoma. Arch Ophthalmol 40:176,1948.

40. Westsmith RA and Abernethy RE: Detachment of retina withuse of diisopropyl fluorophosphate (fluropryl) in treatment ofglaucoma. Arch Ophthalmol 52:779, 1954.

41. Heimann K and Kyrielcis E: Netzhautablosung bei miotischerTherapie. Klin Monatsbl Augenheilkd 156:98, 1970.

42. Lutjcn-Drecoll E, and Kaufman P: Echothiophatc-inducedstructural alterations in the anterior chamber angle of the cyno-molgus monkey. Invest Ophthalmol Vis Sci 9:918, 1979.

43. Feeny-Burns L and Ellersicck MR: Age-related changes in theultrastructure of Bruch's membrane. Am J Ophthalmol100:686, 1985.

44. Hogan MJ and Alvarado J: Studies on the human macula. IV.Aging changes in Bruch's membrane. Arch Ophthalmol77:410.

45. Ishibashi T, Sorgente N, Patterson R. and Ryan SJ: Agingchanges in Bruch's membrane of monkeys. An electron micro-scopic study. Ophthalmologies 192:179, 1986.

46. Leung DYM? Glagov S, and Mathews MB: Cyclic stretchingstimulates synthesis of matrix components by arterial smoothmuscle cells in vitro. Science 191:475. 1976.

47. Gabella G and Yamcy A: Synthesis of collagen by smooth mus-cle in the hypertrophic intestine. QJ Exp Physio! 62:257, 1977.

48. Gabella G: Hypertrophic smooth muscle. V. Collagen andother extracellular materials. Vascularization. Cell Tiss Res235:275. 1984.

49. Imayama S and Bravcrman 1: A hypothetical explanation forthe aging of the skin. Chronologic alteration of the three-di-mensional arrangement of collagen and elastic fibers in con-nective tissue. Am J Pathol 134:1019, 1989.

50. Fisher RF: The force of contraction of the human ciliary mus-cle during accommodation. J Physiol 270:51. 1977.

51. Swegmark G: Studies with impedance cyclography on humanocular accommodation at different ages. Acta Ophthalmol47:1186. 1969.

52. Stievc R: Ubcr den Bau des mcnschlichen Ciliarmuskcls. seinephysiologischen Vcranderungcn wahrend des Lebens undseine Bedeutung fur die Akkommodation. Z Mikrosk AnatForsch55:3, 1949.

53. Rohcn JW: Altersveranderungen im Bereich des vordcrcn Au-gensegments. In Handbuch dcr Gcrontologic, Vol. 3, Platt D,Hockwin O and Mertc H-J, editors. Stuttgart, New York, Gus-tav Fischer Vcrlag, 1989, pp. 68-91.

Documents

Investigative Ophthalmology & Visual Science, Vol. 32, No ... · 1680 INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / April 1991 Vol. 32 Fig. 3. Oblique-tangential section of the posterior