Examination of the anterior uveoscleral pathway in domestic species.

OBJECTIVE: To investigate the uveoscleral (US) pathway in the normal eyes of four domestic spp.: the cat, pig, cow and horse by examining the comparative anatomical structure of anterior US pathway. ANIMALS STUDIED: Four cats, ten pigs, four cows, eight horses. PROCEDURES: Formalin-preserved specimens from anterior uveas of the cat, pig, cow and horse were embedded and serially sectioned sagittally and tangentially and stained with H&E, Masson's trichrome, smooth muscle actin immunolabel, or elastin stain. RESULTS: Spaces between the endings of the outer anterior ciliary body musculature (CBM) formed avenues for the beginning of the US pathway and varied in the amount of extracellular matrix (ECM) material being most developed in the pig. In the cow, other anterior muscle bundles attached the CBM to the sclera concomitant with the presence of an anterior elastic sheath. In the horse, these muscle bundles were connected to branching connective tissue trabeculae within the US pathway that were attached radially to the sclera. In the cat, muscle bundles were more abundant and formed a fine meshwork of trabecular associations with the posterior ICA. Supraciliary development was most pronounced in the horse and least in the pig. CONCLUSION: All species possessed clearly developed and unique US pathways. The outermost muscle bundles of the CBM appeared to have close interaction with the US pathway and the degree of these muscle associations differed with species. The species specific anatomical variations within the US pathway could play a pivotal role in the variability of aqueous outflow along this pathway.

Morphology of the long and short uveal nerves in the human eye.

The aim of this study was to examine the architecture of the uveal nerves in the sclera and suprachoroid of human eyes. Eyes from 17 adult human donors were investigated. The uveal nerves in different regions (retrobulbar, intrascleral, suprachoroidal, pars plana) were prepared and studied by light and electron microscopy. In addition, immunohistochemistry was performed for various neuronal markers. The long uveal nerves showed a characteristic suprachoroidal location with no branches supplying the choroid. It was found that typically they are composed of myelinated (75%) and non-myelinated (25%) nerve fibres. They mainly contain aminergic and sensory nerve fibres. A separate set of cholinergic non-myelinated nerve fibre bundles runs parallel with these long uveal nerves. The short uveal nerves supply the suprachoroidal nerve plexus with approximately 13% of their nerve fibres. The nerves and the branches supplying the choroid appear as mixed nerves containing sympathetic, parasympathetic and sensory axons. This study therefore provides new information about the quantity, type and distribution of myelinated and non-myelinated nerve fibres in the posterior uvea of the human eye.

Innervation of the uvea by galanin and somatostatin immunoreactive axons in macaques and baboons.

The neuropeptide galanin has not been localized previously in the primate uvea, and the neuropeptide somatostatin has not been localized in the uvea of any mammal. Here, the distribution of galanin-like and somatostatin-like immunoreactive axons in the iris, ciliary body and choroid of macaques and baboons using double and triple immunofluorescence labeling techniques and confocal microscopy was reported. In the ciliary body, galanin-like immunoreactive axons innervated blood vessels and the ciliary processes, particularly at their bases. In the iris, the majority of these axons was associated with the loose connective tissue in the stroma. Somatostatin-like immunoreactive axons were found in many of the same areas of the uvea supplied by cholinergic nerves. In the ciliary body, there were labelled axons within the ciliary processes and ciliary muscle. They were also found alongside blood vessels in the ciliary stroma. In the iris, somatostatin-like immunoreactive axons were abundant in the sphincter muscle and less so in the dilator muscle. A unilateral sympathectomy had no effect on the distribution of somatostatin-like or galanin-like immunoreactive axons, and these axons did not contain the sympathetic marker tyrosine hydroxylase. They did not contain the parasympathetic marker choline acetyltransferase, either. The galanin-like immunoreactive axons contained other neuropeptides found in sensory nerves, including calcitonin gene-related peptide, substance P and cholecystokinin. Somatostatin-like immunoreactive axons did not contain any of these sensory neuropeptides or galanin-like immunoreactivity, and they were neither labelled with an antibody to 200kDa neurofilament protein, nor did they bind isolectin-IB(4). Nevertheless, they are likely to be of sensory origin because somatostatin-like immunoreactive perikarya have previously been localized in the trigeminal ganglion of primates. Taken together, these findings indicate galanin and somatostatin are present in two different subsets of sensory axons in primate uvea.

Occurrence of catecholaminergic nerve fibers in the human uveoscleral tissue in conditions of normal and raised intraocular pressure.

The distribution of catecholaminergic nerve fibers (CNF) in human uveoscleral tissue was studied in six human eyes with normal intraocular pressure and in five eyes with increased pressure. The eyes with increased pressure had no visual field alterations and the patients did not have any glaucoma-related optic neuropathies. The amount of norepinephrine in these structures was also analysed. Catecholaminergic nerve fibers were detected by means of fluorescence microscopy and were counted using the quantitative analysis of images. Our results demonstrate that the occurrence of catecholaminergic nerve fibers (expressed in Conventional Units = C.U.) in human uveoscleral tissue is 15.4 +/- 1.6 C.U. in eyes with normal intraocular pressure. In eyes with increased intraocular pressure, these values were 12.2 +/- 1.2 C.U. Moreover, the amount of norepinephrine in tissue homogenates of the same eyes was evaluated and found to be 21.7 +/- 1.3 microg/gr tissue fresh weight of the human uveoscleral tissue in eyes with normal intraocular pressure. This value decreased to 18.8 +/- 1.1 microg/gr tissue fresh weight in the same tissue in conditions of raised intraocular pressure. In these experiments, the small number of eyes examined made it difficult to draw general conclusions. However, the role of human uveoscleral tissue was emphasized by the rich catecholaminergic innervation. A decrease of catecholaminergic nerve fibers and norepinephrine occurs when intraocular pressure is elevated. The modifications of these parameters, involved in the sympathetic control of aqueous humor outflow, may support the hypothesis of a possible relevant role for the human uveoscleral tissue in different pathological conditions.

Light microscopy of uveoscleral drainage routes after gelatine injections into the suprachoroidal space.

PURPOSE: To visualize the suprachoroidal space and to study the morphology of possible uveoscleral drainage routes in the human eye. METHODS: Indian ink stained gelatine was injected directly into the suprachoroidal space of 7 human donor eyes. Those quadrants where gelatine appeared in the episcleral venous network were cut in their entirety in consecutive sections and examined by light microscopy. RESULTS: Gelatine was observed in the connective tissue surrounding scleral vessels and nerves. Gelatine was also traceable in fine, endothelium-lined channels at the inner aspect of the anterior sclera. These channels seemed to originate at the inner surface of the sclera, close behind the scleral spur, and communicate with the intrascleral venous plexus. CONCLUSION: Fluid is drained from the suprachoroidal space through the perivascular and perineural spaces of scleral blood vessels and nerves. In addition, this study indicates that there are preformed channels at the inner aspect of the anterior sclera, capable of draining fluid directly into scleral veins.

Occurrence of cholinergic nerve fibers in the human uveoscleral tissue.

We have studied the cholinergic nerve fibers (CNF) of human uveoscleral tissue in normal and pathological conditions (intraocular pressure: (IOP) of various degrees ) and have performed a quantitative image analysis of CNF with the aim of studying an hypothetical association between IOP and the uveoscleral level of cholinergic neurotransmitter in nerve fiber of the enucleated human eyes. Eleven patients, previously enrolled in our medical protocols are affected by an increase of IOP of various degrees, were subjected to surgical enucleation of one eyeball for traumatic or neoplastic reasons and were the donors of samples of the uveoscleral tissue for CNF demonstration. The samples were also submitted to quantitative image analysis. An enzymatic activity, Acetylcholinesterase (AChe), is present in nerve fibers that innervate the human uveoscleral tissue. The AChe-positive nerve fibers represent about 7.2% of the observed area in normal conditions, while in patients with elevated IOP AChe activity is about 20.3% of the total observed area. The increase of AChe activity in our experiments, all performed under the same conditions, and therefore, with comparable results, may be related with the increase of IOP.

Nitric oxide as a mediator of parasympathetic vasodilation in ocular and extraocular tissues in the rabbit.

PURPOSE: The aim of this study was to investigate in rabbits the relationship between nitric oxide and the noncholinergic vasodilation caused by facial nerve stimulation in the eye and some extraocular tissues. METHODS: Uveal vascular resistance was determined by measuring simultaneously the flow from a cannulated vortex vein with intraocular pressure and arterial blood pressure recordings. The local blood flow in different parts of the eye (iris, ciliary body, choroid, and retina), eyelids, nictitating membrane, Harderian gland, and lacrimal gland was determined using radioactive microspheres. The effects of facial nerve stimulation, at different frequencies, were examined before and after the administration of nitric oxide synthase (NOS) inhibitors. RESULTS: In the experiments with direct determination of uveal blood flow, stimulation of the facial nerve caused a frequency-dependent decrease in uveal vascular resistance, indicating vasodilation. This effect was reduced or abolished by NOS inhibition at low frequencies but was unaffected at high frequencies. Determination of regional blood flow, with radioactive microspheres, showed that the stimulation increased local blood flow in all parts of the uvea. Compared to the nonstimulated control side, stimulation at 2 Hz increased choroidal blood flow by 89% +/- 12% before NOS inhibition and by 45% +/- 10% after NOS inhibition, a difference of 44% +/- 77% (n = 9; P < or = 0.05). Iris and ciliary body vasodilation appeared to be equally reduced. In eyelids, Harderian gland, and lacrimal gland, the vasodilation elicited by stimulation at 2 Hz was abolished almost completely by NOS inhibition. The vasodilation in most of the extraocular tissues was reduced significantly by NOS inhibition at 5 Hz, with only a slight reduction in the choroid, iris, and ciliary body. Retinal blood flow also was significantly increased by facial nerve stimulation at 2 Hz and 5 Hz. The increase in retinal blood flow appeared to be more sensitive to NOS inhibition than the increase in uveal blood flow. CONCLUSIONS: These results suggest that the formation of nitric oxide plays an important role in the uveal, retinal, and extraocular vasodilation brought about by facial nerve stimulation at low frequencies. At high frequencies, other neurotransmitters also seem to be involved.

Distribution and effects of pituitary adenylate cyclase-activating peptide in the rabbit eye.

Pituitary adenylate cyclase-activating peptide (PACAP)-like immunoreactivity was demonstrated by immunocytochemistry together with calcitonin gene-related peptide (CGRP)-like immunoreactivity in small to medium-sized neurons in the trigeminal ganglion and in nerve fibers in the iris, ciliary body, cornea, choroid and sclera of the rabbit eye. The regional distribution of PACAP-27- and PACAP-38-like immunoreactivity in the eye was studied by radioimmunoassay: the highest concentrations were found in the iris sphincter and ciliary body. The distribution pattern resembled that of CGRP-like immunoreactivity, which is a well-known constituent of sensory C-fibre neurons. Intravitreal injection of PACAP-27 or PACAP-38 induced conjunctival hyperemia, swelling of the anterior segment of the eye, miosis and breakdown of the blood-aqueous barrier, manifested as a marked aqueous flare response. Tetrodotoxin pretreatment inhibited the conjunctival hyperemia, the swelling of the anterior segment of the eye, and the miosis but not the aqueous flare response. The concentration of PACAP-like immunoreactivity in the aqueous humor was increased greatly following infrared irradiation of the iris, topical application of formaldehyde to the cornea, or intravitreal injection of endotoxin or bovine serum albumin. Also the concentration of CGRP-like immunoreactivity in the aqueous humor was increased greatly. Both in vivo and in vitro studies showed that capsaicin caused a parallel release of PACAP-like immunoreactivity and CGRP-like immunoreactivity from the uvea. Injection of PACAP-27 and PACAP-38 resulted in the release of CGRP-like immunoreactivity (and PACAP-like immunoreactivity) into the aqueous humor and PACAP-27 and PACAP-38 were also found to evoke tachykinin-mediated contractions of the isolated iris sphincter muscle, indicating that PACAP induces positive feedback on C-fibres. Thus, PACAP is a sensory neuropeptide in the eye. Since the PACAP-induced ocular responses mimicked the symptoms of inflammation, and since the PACAP-like immunoreactivity concentration in the aqueous humor was greatly increased following noxious stimulation, we suggest that it takes part in the inflammatory responses of the rabbit eye.

The 1990 Endre Balazs Lecture. Effects of some neuropeptides on the uvea.

This lecture summarizes studies on the effects of some of the neuropeptides which seem to be present in somatosensory and autonomic nerves in the uvea. Release of these peptides is likely to explain nerve induced effects in the eye which are not due to classical transmitters. Trigeminal nerve fibres in the eye seem to contain substance P (SP), calcitonin gene-related peptide (CGRP), and cholecystokinin (CCK), parasympathetic nerve fibers from the facial nerve seem to contain vasoactive intestinal polypeptide (VIP), and peptide with histidine and isoleucine terminals (PHI), and sympathetic nerves seem to contain neuropeptide Y (NPY). Retrograde trigeminal nerve stimulation in rabbits causes hyperemia, miosis, a breakdown of the blood-aqueous barrier to plasma proteins and a rise in intraocular pressure (IOP). There is release of SP and CGRP or related peptides. The miosis seems to be due to SP and the other effects to CGRP and small amounts of arachidonic acid metabolites released by the peptides. SP has no miotic effect in monkeys and cats. However, CCK is a potent miotic in monkeys and causes contraction of the human pupillary sphincter muscle. It has no such effect in the lower species. The effect of CCK in primates seems to derive from the presence of CCK receptors of the A-type on the pupillary sphincter muscle, and can be blocked by lorglumide. Miosis can be produced in cats by the peptide endothelin; this effect is due to release of arachidonic acid metabolites. Facial nerve stimulation causes vasodilation in the uvea of rabbits, cats and monkeys. The effect cannot be abolished by muscarinic blocking agents.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcitonin gene-related peptide (CGRP) immunoreactive sensory nerves in the human and guinea pig uvea and cornea.

The presence of calcitonin gene-related peptide (CGRP) immunoreactive nerves in the uvea and cornea of human and guinea pig eyes was evaluated using immunohistochemical techniques. CGRP immunoreactivity was found in thin, varicose nerve fibers in both species. Most of the fibres were localized in the ciliary body, and were mainly associated with blood vessels. In the human ciliary body, a moderate number of CGRP immunoreactive nerves were also seen in the ciliary muscle. In the iris and cornea, CGRP immunoreactive fibres were relatively uncommon. In the iris, they were mostly found associated with blood vessels, while in the cornea they were seen sub-epithelially or as free nerve endings in the epithelium. In the trigeminal ganglion, small sized ganglion cells displayed CGRP immunoreactivity. About 40% of all ganglion cells were immunoreactive nerves in the guinea pig, while sympathetic denervation did not change the staining pattern of CGRP immunoreactivity. The present findings, together with previous physiological data, suggest that CGRP might play a role in the regulation of the blood flow, aqueous humour dynamics, and neurogenic inflammation, not only in experimental animals but also in man.

Nerve fibers showing immunoreactivities for thyrosine hydroxylase and dopamine-beta-hydroxylase re-appear in the guinea pig uvea after sympathectomy.

After sympathectomy we have studied the re-appearance of nerve fibers showing catecholaminergic characteristics in the uvea of the guinea pig. Immunoreactivities for two catecholamine symthetizing enzymes, tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH), were used as markers. Both TH-like and DBH-like immunoreactive nerve fibers disappeared after the extirpation of ipsilateral superior cervical ganglion. In the choroid the TH-like and DBH-like immunoreactive nerve fibers re-appeared within 2 weeks. In the iris and the ciliary body both of these types of immunoreactive nerve fibers re-appeared 10 weeks after the denervation. The morphological appearance of these re-appearing nerve fibers was not similar to those in the non-denervated uvea.

Vasoactive intestinal polypeptide and the ocular innervation.

The indirect immunofluorescence technique with antisera to vasoactive intestinal polypeptide (VIP) stains peripheral nerve fibers in both the anterior segment and the posterior segment of rat, guinea pig, cat, and rhesus monkey eyes. While immunoreactive corneal nerves are lacking, all four species have a prominent innervation of the superficial limbal blood vessels. The aqueous humor outflow apparatus of the rat, guinea pig, and cat, but not the monkey, contain VIP-like immunoreactive nerves. All four animals have immunoreactive iris nerve fibers, tending either to surround large blood vessels or to lie as free stromal nerves. Only in the cat are immunoreactive nerve fibers seen within the iris muscles. A modest number of VIP-like immunoreactive nerves are present in the ciliary body of all four animals; immunoreactive nerve fibers within the ciliary processes occur only in the rat and guinea pig. VIP-like immunoreactive nerves are found in the choroid of all four animals. An association of immunoreactive nerve fibers to uveal melanocytes also is apparent. The present findings expand several prior immunohistochemical studies of mammalian eyes in which the VIP-like immunoreactive nerves to the choroid was emphasized.

Sympathetic uveitis 66 years after injury.

We report a case of sympathetic uveitis occurring 66 years after perforating trauma to the other eye. To our knowledge this is the longest interval ever cited in the literature.

Sensory supply of the anterior uvea: a light and electron microscope study.

We have studied the ultrastructural features of sensory nerve fibers in the ciliary body and the iris and their parent trigeminal ganglion cells, using intra-axonally transported horseradish peroxidase as a tracer. Unmyelinated nerve fibers of ipsilateral trigeminal origin were found in stroma of both the ciliary body and the iris. Most appeared in bundles of nerve fibers; but some were found singly. Two distinct types of varicosities were found, one containing both a few irregularly shaped vesicles and mitochondria, the other containing mitochondria only. The thin profiles of the axons between these varicosities or endings contained neurotubules and filaments. The trigeminal ganglion cells supplying the anterior eye were of a relatively small size (15-50 micron in diameter) and confined to the anteromedial part of the ipsilateral ganglion. These cells could not be classified according to the arrangement of their subcellular organelles. The morphological characteristics of the sensory nerve fibers and ganglion cells supplying the anterior uvea are consistent with the view that pain may be the predominant sensation mediated by these neurons.

Pancreatic polypeptide-like immunoreactive nerves in the guinea pig eye.

The indirect immunofluorescence technique with antisera either to avian pancreatic polypeptide or to bovine pancreatic polypeptide stains nerve fibers in the guinea pig eye. In all regions of the uvea, immunoreactive fibers are present around large blood vessels; an association of immunoreactive nerve fibers to melanocytes is seen. Immunoreactive nerves are found throughout the choroid, including the choriocapillaris. In the ciliary body, they are seen in individual ciliary processes. The iris dilator muscle and, to a lesser degree, its sphincter are innervated. The chamber angle of the anterior segment contains immunoreactive nerve fibers, but convincing innervation to the cornea is lacking. No retinal cells stain. With some exceptions, the distribution of peripheral nerve fibers parallels that of the adrenergic innervation. Appropriate controls are negative.

Neuropeptide Y-immunoreactive nerves in the uvea of guinea pig and rat.

Neuropeptide Y (NPY)-immunoreactivity has been localized to a rich network of fibres in the uvea of guinea pig and rat. In the iris, NPY-immunoreactive nerves were present in iridial smooth muscle and around blood vessels; similarly throughout the choroid a dense network of NPY-immunoreactive fibres was found surrounding the choroidal vascularization. Following sympathectomy a marked decrease of NPY-immunoreactivity, in particular in the iris and less marked in the choroid, was noted, indicating that these fibres may originate at least in part from neuronal cell bodies in the superior cervical ganglion. The close relationship of the NPY-immunoreactive fibres to blood vessels in both anterior and posterior uvea is consistent with the established vasoconstrictor action of this peptide.

Mapping, quantitative distribution and origin of substance p- and VIP-containing nerves in the uvea of guinea pig eye.

VIP- and substance P-like immunoreactivities were found in considerable concentrations (VIP: 17.3 +/- 4.8 pmol/g, mean +/- SEM; substance P:11.1 +/- 1.8 pmol/g) in the uveal portion of the guinea pig eye. Immunocytochemistry localised these two regulatory peptides to nerve fibres found principally in a plexus in the iris (substance P) and in an extensive network surrounding the blood vessels of the choroid (VIP). A remarkable anatomical demarcation of the two types of peptide-containing nerves was established by the staining of substance P-containing nerves, which stops at the level of the ciliary body. This uveal area is known to be involved in the ocular responses to nociceptive stimuli. At the ultrastructural level, immunoreactivity for both peptides was localised to distinct subpopulations of p-type nerves, distinguishable by the size of their large dense-cored vesicles. Those immunoreactive for VIP were significantly larger (p less than 0.0005) than those immunoreactive for substance P (95 +/- 7 nm and 82 +/- 9 nm respectively; mean +/- SD). Interruption of the trigeminal pathway produced a remarkable decrease of substance P immunoreactivity in the anterior portion of the uvea (9.1 +/- 1.5 pmol/g, mean +/- SEM, control; 5.3 +/- 1.3 pmol/g, denervated), but not of VIP immunoreactivity in the choroid. Following colchicine treatment, VIP-immunoreactive neuronal cell bodies were localised in the choroid. The separate anatomical localisations and distributions of the two uveal peptides appear to be related to their different origins and functional roles in the response of the eye to noxious stimuli.

Vasoactive intestinal peptide nerves in ocular and orbital structures of the cat.

Vasoactive intestinal polypeptide (VIP), a neuronal peptide of ubiquitous occurrence in the body, is known to have strong vasodilatory effects and to promote secretion from many exocrine glands. Nerves displaying VIP immunoreactivity (VIP nerves) were detected in several orbital structures of the cat. Such nerves were numerous in the lacrimal glands and somewhat less numerous in the Harderian glands and the tarsal glands. The nerves surrounded glandular acini and small blood vessels. Intraocularly, VIP nerves were seen in the ciliary processes, in the posterior third of the ciliary muscle, and around small to medium-sized blood vessels in the posterior uvea. VIP nerve fibers were absent from vessels in the anterior uvea. This distribution may explain why intracranial stimulation in the oculomotor nerve exit region dilates the vessels of the choroid but not those of the iris. A large number of VIP-immunoreactive nerve cell bodies were observed in the pterygopalatine ganglion. Extirpation of this ganglion resulted in the disappearance of VIP nerves from the intraocular structures and from the lacrimal and Harderian glands. Removal of the superior cervical ganglion and the ciliary ganglion did not affect the VIP nerve supply. The results suggest that the VIP nerves originate in the pterygopalatine ganglion.