Investigative Ophthalmology & Visual Science, Vol. 30, No. 9, September 1989 Copyright © Association for Research in Vision and Ophthalmology Elastic Fibers in the Rat Exorbital Lacrimal Gland Duct System Mortimer Lorber In the rat, each paired subcutaneous exorbital lacrimal gland overlies the retromandibular area. The fibrous cord containing its ducts passes over the masticatory muscles to the temporal canthus. Thus, the lacrimal system must accomodate both jaw and lid movements. To see whether elastic fibers exist to modulate alterations in tension caused by such movements, light and electron micrographs were made of its duct system. The innermost elastic fibers are connected by elaunin fibrils and oxytalan microfibrils to the lamina densa, particularly near the hemidesmosomes. The innermost elastic fibers appear to be longitudinal and about 0.2 nm from the lamina densa. Circumferential fibers exist about 0.8-3.2 nm from that structure. More peripheral fibers of both orientations also exist. Light micros- copy of the extraglandular duct demonstrated circumferential fibers near the basement membrane and longitudinal and angular elastic fibers amid the collagenous layers. Some of the longitudinal fibers assume a loose cross-weave. Intraglandularly, as duct size diminishes elastic fibers progressively decrease in number and size until the smallest ducts have none. Thus, an elastic gradient exists. It is believed that recoil of the angular elastic fibers aids distension of the large and medium ducts when secretion is great and that recoil of the circumferential ones permits those duct diameters to diminish when secretion does. The longitudinal elastic fibers would allow all but the smallest ducts to recoil from the stretching of much of the exorbital lacrimal duct system accompanying blinking and other facial movements. Invest Ophthalmol Vis Sci 30:2002-2011,1989 Rat exorbital lacrimal glands are paired, shield- organ itself that might modulate dimensional shaped, subcutaneous structures overlying the front changes was performed. Their presence was antici- of the parotid gland, the retromandibular area and pated because in the embryo the lacrimal gland de- the rear portion of the body of the mandible. Venable velops as an evagination of the conjunctiva.4 The lat- and Griffin1 noted that they are oriented perpendicu- ter is known to contain elastic tissue,5'6 so it should larly to the zygomatic arch, resting on the superficial not be surprising if the epithelial outpouching that muscles of mastication. They lie slightly below and becomes the duct and eventually the gland carries just in front of each ear.2 Because these serous, tu- with it some of the collagen and elastic fibers of the bulo-acinar glands3 are the largest components of the connective tissue surrounding the bud. This presum- rodent lacrimal system, presumably they are the ably explains not only the present observations but major contributors to the secretions moistening the those made by Sundwall in 19167 that elastic fibers surface of the ipsilateral eye. Each has a gross struc- exist in the lacrimal gland of the ox. In 1897, Fuma- ture appearing to be its duct that extends between the galli8 described an intricate network of what he upper part of the organ near its anterior border and termed elastic fibers throughout the human lacrimal the conjunctiva near the temporal canthus. gland. Although the internal elastic membrane of ar- Because of the slight displacement of the exorbital teries was stained, his illustrations and description lacrimal gland as the mandible moves and the need of indicate that he often deemed reticulin fibers to be its duct to alter its length with eyelid movements, that elastic fibers in addition to those meriting such classi- is, stretching as the lids blink and recoiling as they fication. open, a search for elastic fibers in the duct and the Materials and Methods From the Department of Physiology and Biophysics, George- One or both exorbital lacrimal glands were re- town University School of Medicine, Washington, DC. moved from six adult rats of both sexes following Submitted for publication: November 14, 1988; accepted March anesthesia with intraperitoneally administered so- 13, 1989. 9 Reprint requests: Mortimer Lorber, DMD, MD, Department of dium pentobarbital. For light microscopic studies, Physiology and Biophysics, Georgetown University School of Med- eight glands, often with a portion of their gross duct, icine, 3900 Reservoir Road, N.W., Washington, DC 20007. were excised from five animals and fixed in phos- 2002 Downloaded from iovs.arvojournals.org on 10/02/2021 No 9 RAT EXORBITAL LACRIMAL DUCT ELASTIC FIBERS / Lorber 2003 Fig. 1. Extraglandular portion of rat lacrimal duct stained for elastic tissue with orcein. (a) Cross-section of the cord which grossly appears to be the duct midway in its length. Two dissimilar ducts with precipitated luminal secretion are evident. At least five arterioles and one large venule (V) are present. Bar = 100 /im. (b) Higher magnification of bottom of the upper duct in (a). An occasional circumferential elastic fiber (arrows) immediately underlies the basement membrane. Numerous transected longitudinal ones exist more peripherally in several layers. Bar = 10 iim. phate-buffered neutral formalin, dehydrated and em- lacrimal gland indicates that at least part of the struc- bedded in paraffin. Sectioning was done at 5 nm fol- ture contains two ducts of dissimilar size and shape lowed by hydration and staining with a Weigert's res- (Fig. la). orcin-fuchsin stain or a modified Tanzer's stain of Most elastin lies near the duct epithelium. Immedi- 0.4% orcein in acidified ethanol with Van Gieson ately subepithelially, a layer of delicate circumferen- counterstaining. tial elastic fibers about 0.2-0.5 urn in diameter is seen For electron microscopy, part of the gross extra- (Fig. lb). Many larger, longitudinal elastic fibers, up glandular duct of one adult rat of each sex was fixed to 1.2 nm in diameter, exist more peripherally in a for 1 hr in 5% glutaraldehyde and 4% paraformalde- number of layers (Figs, lb, 2a). Some angular elastic hyde in 0.2 M phosphate buffer, pH 7.4. The tissue fibers that extend diagonally or somewhat perpendic- was postfixed for one hour in 1% osmium tetroxide in ularly are more peripheral (Fig. 2b). They are about 0.2 M s-collidine buffer at pH 7.4, a modified Kar- 0.4-0.8 /jm in diameter. Because they do not encom- novsky procedure.10 En bloc staining with 2% pass the duct and are often not truly perpendicular to aqueous uranyl acetate was followed by dehydration. the basement membrane, they have not been termed Transversely and longitudinally oriented pieces were radial fibers. then embedded in epoxy resin. Survey sections (1 There is also a loose weave of delicate elastic fibers, nm) were stained with 0.1% toluidine blue in 2% so- about 0.2-0.4 ^m in diameter, that is occasionally dium borate. Selected areas were sectioned at 0.08 seen close to the base of the epithelium (Fig. 2c). fim, placed on copper grids and exposed to 2% uranyl These connect with the surrounding thicker and acetate in 25% ethanol for 15 min followed by 0.04 M more numerous longitudinal elastic fibers (Fig. 2c, d). lead citrate for one to 2 min." The thin sections were Ducts in which a loose elastic weave is evident may examined using a JEOL transmission electron micro- be more relaxed than those that do not display that scope (Tokyo, Japan). architecture. Relaxation is indicated by unusually Light microscopic measurements of dimensions great undulation of collagen fibers (Fig. 2c). Whether and distances were made using an ocular reticle that this diminished tension existed in situ or developed had been calibrated by an objective micrometer. subsequently is not known. These studies accord with the ARVO Resolution Within the gland, between the largest ducts whose on the Use of Animals in Research. lumens have a maximum dimension of about 45-75 urn and the smallest ducts, the elastic fibers progres- Results sively diminish in number and often in thickness. Light Microscopy The main duct and its largest branches have a rela- Histological examination of what is grossly a single tively few, delicate circumferential fibers, often about cord appearing to be the duct of the rat exorbital 0.3-0.4 fim in diameter, usually appearing in a single Downloaded from iovs.arvojournals.org on 10/02/2021 2004 INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / September 1989 Vol. 30 Fig. 2. Extraglandular portion of rat lacrimal duct to demonstrate elastic tissue, (a), (b) and (d) are stained with Weigert's, (c) is stained with orcein. (a) Wall of transected duct showing a few delicate, dark, circumferential elastic fibers (arrows) between the epithelium and the innermost circumferential collagen fibers. Most elastic fibers are transected longitudinally as are the large collagen fibers in the center. Bar = 5 f*m. (b) The transected duct has some circumferential fibers (arrow) both beneath the epithelial cells and at a distance from them (arrowhead). Transected longitudinal elastic fibers and more peripheral, thicker, angular elastic fibers (A) are near the upper left of the duct. Bar = 10 Mm. (c) Longitudinal section grazing the base of the epithelium to reveal a loose cross-weave (arrowheads) of elastic fibers through which the bases of the epithelial cells are seen. The undulating collagen suggests the tissue is relaxed. Many longitudinal elastic fibers (arrows) exist, particularly beneath the duct where the thick collagen fibers stain prominently. Some of those elastic fibers may connect with those in the cross-weave. To the left of the left arrow are several transected circumferential elastic fibers. Bar = 10 /urn. (d) About the longitudinally sectioned duct whose lumen (L) is visible very long elastic fibers are asymmetrically arranged, ie, those above the duct are fewer and often thicker than those beneath it.