Type-specific photoreceptor loss in pigeons after disruption of parasympathetic control of choroidal blood flow by the medial subdivision of the nucleus of Edinger-Westphal.

The medial part of the nucleus of Edinger-Westphal (EWM) in birds mediates light-regulated adaptive increases in choroidal blood flow (ChBF). We sought to characterize the effect of loss of EWM-mediated ChBF regulation on photoreceptor health in pigeons housed in either moderate intensity diurnal or constant light (CL). Photoreceptor abundance following complete EWM destruction was compared to that following a lesion in the pupil control circuit (as a control for spread of EWM lesions to the nearby pupil-controlling lateral EW) or following no EW damage. Birds were housed post-lesion in a 12 h 400 lux light/12 h dark light cycle for up to 16.5 months, or in constant 400 lux light for up to 3 weeks. Paraformaldehyde-glutaraldehyde fixed eyes were embedded in plastic, sectioned, slide-mounted, and stained with toluidine blue/azure II. Blinded analysis of photoreceptor outer segment abundance was performed, with outer segment types distinguished by oil droplet tint and laminar position. Brains were examined histologically to assess lesion accuracy. Disruption of pupil control had no adverse effect on photoreceptor outer segment abundance in either diurnal light or CL, but EWM destruction led to 50-60% loss of blue/violet cone outer segments in both light conditions, and a 42% loss of principal cone outer segments in CL. The findings indicate that adaptive regulation of ChBF by the EWM circuit plays a role in maintaining photoreceptor health and mitigates the harmful effect of light on photoreceptors, especially short wavelength-sensitive cone photoreceptors.

Angioarchitecture and innervation of the primate anterior episclera.

PURPOSE: To investigate the primate episcleral vasculature and its innervation with respect to morphological specializations. METHODS: Serial sections of the anterior episclera of 8 monkey eyes and 20 human eyes were investigated enzyme- and immunohistochemically using antibodies against smooth-muscle alpha-actin (SMA), neurofilament, synaptophysin, substance P (SP), calcitonin gene-related peptide (CGRP), vesicular acetylcholine transporter (VACHT), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), tyrosine hydroxylase (TH), vesicular monoamine transporter II (VMAT II), as well as the NADPH-diaphorase reaction. Arteriovenous anastomoses (AVA) were quantified. RESULTS: All episcleral vessels including veins showed intense staining for SMA. Capillary loops were only seen in the limbal arcades, not in the episclera itself. Instead, AVA connected the episcleral arteries with the veins, which formed an interlacing vascular network. In the monkey episclera, 4-6/mm2 AVA were found; in the human episclera, 0.5-1/mm2. Numerous nerve endings staining for NADPHd (NADPHdiaphorase) and TH surrounded all episcleral vessels including anastomoses and veins. NPY, VIP, and VACHT-immunoreactive (IR) nerve terminals were less numerous. CGRP and SP-IR terminals were seen both at the vessels and in the intervascular connective tissue. CONCLUSIONS: The episcleral vasculature shows a specialized morphology with absence of capillaries, numerous arteriovenous anastomoses, a muscle-rich venous network, and intense innervation by vasodilative and vasoconstrictive nerves. This might allow regulation of blood flow and volume in the episcleral vessels and Voigt's capillaries for thermoregulation and modulation of episcleral venous pressure and thereby outflow facility.

Biphasic neurogenic vasodilatation in the bovine intraocular long posterior ciliary artery: involvement of nitric oxide and an additional unidentified neurotransmitter.

We have investigated the neurogenic factors inducing relaxation in the intraocular segment of the bovine long posterior ciliary artery. In precontracted vessels, electrical field stimulation (EFS, 0.5-128 Hz, 10 s trains) in the presence of guanethidine (30 microM) evoked biphasic relaxation: optimal relaxation for the first and second components occurred at 10 and 50 s, respectively. The first component, but not the second, was abolished by L-NAME (100 microM) or ODQ (3 microM). Relaxation to exogenous CGRP (0.1-300 nM) was inhibited by the CGRP antagonist, CGRP(8-37) (1-5 microM), but neither component of neurogenic relaxation was affected. Preincubation with the sensory nerve excitotoxin, capsaicin (1 microM), had no effect on either the first or second components of neurogenic relaxation. Substance P (0.1 nM-0.1 microM) induced relaxation, but rapid and complete desensitisation occurred within minutes. Neither desensitisation to substance P (0.1 microM) nor incubation with the NK(1) antagonist, L-733,060 (0.3 microM), had any effect on the first or second components of neurogenic relaxation.VIP (0.1 nM-0.3 microM) induced relaxation and this was followed by substantial desensitisation. Neither desensitisation to VIP (0.6 microM) nor treatment with the protease, alpha-chymotrypsin (10 U ml(-1)), had any effect on the first or second components of neurogenic relaxation. The results indicate that nitric oxide mediates the first component of neurogenic relaxation in the bovine intraocular ciliary artery. The neurotransmitter mediating the second component remains to be determined but is unlikely to be CGRP, substance P or VIP.

Vasoactive intestinal and calcitonin gene-related peptides, tyrosine hydroxylase and nitrergic markers in the innervation of the rat central retinal artery.

The vascular supply of the optic nerve has been studied with different methods including corrosion casts both in humans and in other mammals. In man, primates and some other mammals, such as the rat, a distinct central retinal artery accompanies the optic nerve, and runs through the lamina cribosa to reach the optic nerve head. Similarities between human and rat central retinal artery could serve to understand changes in the autonomic perivascular innervation in glaucoma using the rat as an animal model. Nitric oxide, calcitonin gene-related peptide, neuropeptide Y, substance P and vasoactive intestinal peptide have been identified around the monkey central retina artery. Innervation of the rat central artery, however, has not been described in detail. Using immuno- and histochemical methods, the present study investigates the peptidergic, adrenergic and nitrergic innervation of the rat posterior ciliary artery as well as the central retina artery. Numerous nitric oxide positive nerve fibers were visualized posterior and anterior to the lamina cribosa of the optic nerve. They colocalized with NADPH-diaphorase positive fibers, which could also be observed in two of six specimens studied at the level of the optic nerve head. Calcitonin gene-related peptide, tyrosine hydroxylase, and VIP positive fibers were also observed surrounding the vessels of the rat optic nerve. The presence of neuronal nitric oxide/NADPH-diaphorase and vasoactive intestinal peptide positive nerve fibers surrounding the posterior ciliary and central retinal arteries indicates a vasodilator effect in the rat optic nerve. Tyrosine hydroxylase positive innervation indicates the presence of sympathetic activity, and calcitonin gene-related peptide positive fibers indicate sensory innervation by trigeminal primary efferents.

Neurogenic vasoconstriction as affected by cholinergic and nitroxidergic nerves in dog ciliary and ophthalmic arteries.

PURPOSE: To determine the involvement of noradrenergic and other vasoconstrictor nerves in the contraction of ocular arteries and the modification by cholinergic and nitroxidergic nerves of vasoconstrictor nerve function. METHODS: Changes in isometric tension were recorded in helical strips of the canine posterior ciliary and external ophthalmic arteries denuded of the endothelium, which were stimulated by transmurally applied electrical pulses (5 Hz). Vasoconstrictor mediators were analyzed by pharmacological antagonists, such as prazosin, alpha,beta-methylene ATP, a P2alpha-purinoceptor antagonist, and BIBP3226, a neuropeptide Y receptor antagonist. RESULTS: Transmural electrical stimulation produced contractions that were potentiated by N(G)-nitro-L-arginine (L-NA), a nitric oxide (NO) synthase inhibitor. The contraction was partially inhibited by prazosin and abolished by combined treatment with alpha,beta-methylene ATP but was not influenced by BIBP3226. Stimulation-induced contraction was attenuated by physostigmine and potentiated by atropine. Contractions induced by exogenous ATP were reversed to relaxations by alpha,beta-methylene ATP. In the strips treated with L-NA, prazosin, and alpha,beta-methylene ATP, the addition of L-arginine elicited relaxations by nerve stimulation. The ATP-induced relaxation was attenuated by aminophylline, whereas neurogenic relaxation was unaffected. CONCLUSIONS: Ciliary and ophthalmic arterial contractions by nerve stimulation are mediated by norepinephrine and ATP, which stimulate alpha1-adrenoceptor and P2X purinoceptor, respectively. ATP from the nerve is unlikely involved in vasodilatation. Acetylcholine derived from the nerve impairs the neurogenic contraction, possibly by interfering with the release of vasoconstrictor transmitters, and neurogenic NO also inhibits the contraction postjunctionally by physiological antagonism.

Analysis of blood flow in the long posterior ciliary artery of the cat.

PURPOSE: Experiments were undertaken to use a new technique for direct on-line measurement of blood flow in the long posterior ciliary artery (LPCA) in cats and to evaluate possible physiological mechanisms controlling blood flow in the vascular beds perfused by this artery. METHODS: Blood flow in the temporal LPCA was measured on a continuous basis using ultrasonic flowmetry in anesthetized cats. Effects of acute sectioning of the sympathetic nerve and changes in LPCA and cerebral blood flows in response to altered levels of inspired CO2 and O2 were tested in some animals. In others, the presence of vascular autoregulatory mechanisms in response to stepwise elevations of intraocular pressure was studied. RESULTS: Blood flow in the temporal LPCA averaged 0.58+/-0.03 ml/min in 45 cats anesthetized with pentobarbital. Basal LPCA blood flow was not altered by acute sectioning of the sympathetic nerve or by changes in low levels of inspired CO2 and O2, although 10% CO2 caused a modest increase. Stepwise elevations of intraocular pressure resulted in comparable stepwise decreases of LPCA blood flow, with perfusion pressure declining in a linear manner throughout the perfusion-pressure range. CONCLUSIONS: Ultrasonic flowmetry seems to be a useful tool for continuous on-line measurement of LPCA blood flow in the cat eye. Blood flow to vascular beds perfused by this artery does not seem to be under sympathetic neural control and is refractory to modest alterations of blood gas levels of CO2 and O2. Blood vessels perfused by the LPCA show no clear autoregulatory mechanisms.

Cholinergic nerve function in monkey ciliary arteries innervated by nitroxidergic nerve.

We sought to determine the control of ciliary arterial tone by neurogenic acetylcholine (ACh) acting directly on smooth muscle and in conjunction with vasodilator nerves. Isolated posterior ciliary arteries from monkeys responded to ACh (10(-8)-10(-5) M) with dose-related contractions, which were endothelium independent. The response was not affected by cyclooxygenase inhibitors but was abolished by atropine. Relaxations induced at 10(-4) M ACh in the atropine-treated arterial strips were abolished by hexamethonium and NG-nitro-L-arginine (L-NNA), and L-arginine (L-Arg) reversed the response suppressed by L-NNA. Similar results were also obtained on the nicotine (10(-4) M)-induced relaxation. Contractions due to transmural electrical stimulation in the endothelium-denuded strips treated with L-NNA were potentiated by physostigmine and depressed by atropine; the remaining contraction in the presence of atropine was abolished by prazosin. Relaxations associated with electrical stimulation, sensitive to tetrodotoxin, were abolished or reversed to contractions by L-NNA and restored by L-Arg. Stimulation-induced relaxation was attenuated by exogenous ACh and physostigmine and was potentiated by atropine. ACh did not affect the relaxation caused by nitric oxide (NO). Nerve fibers and bundles containing NADPH diaphorase and acetylcholinesterase were histologically demonstrated in the adventitia of ciliary arteries. We conclude that 1) endogenous and exogenous ACh contracts monkey ciliary arteries by acting on muscarinic receptors in smooth muscle cell membranes, 2) vasodilatation elicited by nerve stimulation with electrical pulses or nicotine is mediated by NO synthesized from L-Arg, 3) neurogenic ACh seems to interfere with the nitroxidergic nerve function by acting on prejunctional muscarinic receptors, and 4) high concentrations of ACh stimulate nicotinic receptors in vasodilator nerve terminals and promote the synthesis and/or release of NO.

Modulation by neurogenic acetylcholine of nitroxidergic nerve function in porcine ciliary arteries.

PURPOSE: To determine whether nitroxidergic, cholinergic, and vasoactive intestinal polypeptide (VIP)-mediated nerves participate in the regulation of porcine ciliary arterial tone and to analyze the mechanisms underlying the neuronal interaction. METHODS: Changes in isometric tension were recorded in helical strips of the arteries, which were stimulated by transmurally applied electrical pulses or nicotine. The presence of perivascular nerve fibers containing reduced nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase, acetylcholinesterase, and VIP immunoreactivity were determined histologically. RESULTS: Transmural electrical stimulation (2, 5, and 20 Hz) and nicotine produced a relaxation of the arterial strips denuded of the endothelium and contracted with prostaglandin F2alpha. The response was not influenced by timolol but was abolished by oxyhemoglobin and methylene blue. N(G)-nitro-L-arginine, a nitric oxide (NO) synthase inhibitor, abolished the neurogenic relaxation, and L-arginine restored the response. Physostigmine inhibited, but atropine potentiated, the neurogenic response. The relaxation was attenuated by acetylcholine but was not influenced by VIP. There were nerve fibers and bundles containing NADPH diaphorase, acetylcholinesterase, and VIP immunoreactivity in the adventitia of ciliary arteries. CONCLUSIONS: Porcine ciliary arteries are innervated by NO synthase-containing nerves that liberate NO, possibly as a neurotransmitter on excitation to produce muscular relaxation. Nitroxidergic nerve function is inhibited by acetylcholine released from cholinergic nerve, possibly because of impaired production or release of NO. VIP does not seem to function as a neurotransmitter or a modulator.