Alice M. McGlinn

Nicotinic Acetylcholine Receptor Subunits in Rhesus Monkey Retina

PURPOSE The purpose of this study was to detect and establish the cellular localizations of nicotinic acetylcholine receptor (nAChR) subunits in Rhesus monkey retina. METHODS Retinas were dissected from the eyes of monkeys killed after unrelated experiments. RNA was extracted and analyzed by RT-PCR, using primers designed against human sequences of alpha3-alpha7, alpha9, and beta2-beta4 nAChR subunits. The RT-PCR products were separated by gel electrophoresis and sequenced. Frozen sections of postmortem fixed monkey eyes were immunolabeled with well-characterized and specific monoclonal antibodies against the alpha3, alpha4, alpha6, alpha7, beta2, or beta4 nAChR subunits and visualized with fluorescence labeling. RESULTS Products of the predicted size for the alpha3-alpha7, alpha9, and beta2-beta4 nAChR subunits were detected by RT-PCR in Rhesus monkey retina. Homology between transcripts from monkey retina and human nucleotide sequences ranged from 93 to 99%. Immunohistochemical studies demonstrated that neurons in various cell layers of monkey retina expressed alpha3, alpha4, alpha7, or beta2 nAChR subunits and cells with the morphology of microglia were immunoreactive for the alpha6 or beta4 nAChR subunits. CONCLUSIONS nAChR subunits are expressed in the monkey retina and localize to diverse retinal neurons as well as putative microglia. Besides mediating visual processing, retinal nAChRs may influence refractive development and ocular pathologies such as neovascularization.

Image Defocus and Altered Retinal Gene Expression in Chick: Clues to the Pathogenesis of Ametropia

PURPOSE Because of the retinas role in refractive development, this study was conducted to analyze the retinal transcriptome in chicks wearing a spectacle lens, a well-established means of inducing refractive errors, to identify gene expression alterations and to develop novel mechanistic hypotheses about refractive development. METHODS One-week-old white Leghorn chicks wore a unilateral spectacle lens of +15 or -15 D for 6 hours or 3 days. With total RNA from the retina/(retinal pigment epithelium, RPE), chicken gene microarrays were used to compare gene expression levels between lens-wearing and contralateral control eyes (n = 6 chicks for each condition). Normalized microarray signal intensities were evaluated by analysis of variance, using a false discovery rate of <10% as the statistical criterion. Selected differentially expressed genes were validated by qPCR. RESULTS Very few retina/RPE transcripts were differentially expressed after plus lens wear. In contrast, approximately 1300 transcripts were differentially expressed under each of the minus lens conditions, with minimal overlap. For each condition, low fold-changes typified the altered transcriptome. Differentially regulated genes under the minus lens conditions included many potentially informative signaling molecules and genes whose protein products have roles in intrinsic retinal circadian rhythms. CONCLUSIONS Plus or minus lens wear induce markedly different, not opposite, alterations in retina/RPE gene expression. The initial retinal responses to defocus are quite different from those when the eye growth patterns are well established, suggesting that different mechanisms govern the initiation and persistence or progression of refractive errors. The gene lists identify promising signaling candidates and regulatory pathways for future study, including a potential role for circadian rhythms in refractive development.

Peptide immunoreactivity of the ciliary ganglion and its accessory cells in the rat

By means of immunohistochemistry, calcitonin gene-related peptide, Leu-enkephalin and neuropeptide Y localize to rat ciliary and accessory ganglion cells. The proportion of ciliary and accessory neurons immunoreactive to each peptide is provided and compared to previous data for vasoactive intestinal peptide. These findings indicate considerable neurochemical complexity for a parasympathetic ganglion with a small cell population.

Increase of basal cell membrane area of the retinal pigment epithelium in experimental diabetes.

Stereological analysis of electron micrographs of the pigment epithelium of rats with drug-induced diabetes demonstrated an increase of plasma membrane surface area at the basal aspect of the cells. In none of the diabetic animals examined was there any evidence of breakdown of the blood-retinal barrier to the protein tracer, horseradish peroxidase. Statistically significant increases in basal plasma membrane length and surface density (surface area per unit cell volume) were measured in both streptozotocin and alloxan-injected rats after four weeks of diabetes. When hyperglycemia in streptozotocin-injected rats was promptly reversed by transplantation of normal pancreatic islets, the increase of membrane surface area did not occur. We conclude, therefore, that increased basal surface area of pigment epithelial cells is related to the diabetic condition rather than to a toxic action of the diabetogenic agents. Furthermore, increased membrane surface area was present in streptozotocin-diabetic rats killed after six months of diabetes indicating that the structural change is relatively stable. Relation of basal membrane alteration in the pigment epithelium to any functional disturbance of the barrier cell layer or of the retina in diabetes remains to be established.

Brain natriuretic peptide-immunoreactive nerves in the porcine eye.

Using immunohistochemistry, brain natriuretic peptide (BNP)-like immunoreactive (-LI) nerves localize to the anterior segment of the porcine eye. BNP-LI nerve fibers are visualized mainly in the aqueous outflow pathway, ciliary processes and anterior ciliary muscle. A small number of immunoreactive nerve fibers occur in the anterior iris stroma and in relation to the iris muscles and in the cornea. Given the known ocular effects of atrial natriuretic peptide (ANP) and that BNP and ANP seem to share common receptors, these results suggest an ocular role for peptides of this class independent of delivery by the systemic circulation.

Development of Experimental Myopia in Chicks in a Natural Environment.

Purpose The hypothesis that outdoor exposure might protect against myopia has generated much interest, although available data find only modest clinical efficacy. We tested the effect of outdoor rearing on form-deprivation myopia in chicks, a myopia model markedly inhibited by high-intensity indoor laboratory lighting. Methods Unilaterally goggled cohorts of White Leghorn chicks were maintained in a species-appropriate, outdoor rural setting during daylight hours to the extent permitted by weather. Control chicks were reared indoors with incandescent lighting. Besides ocular refraction and ultrasound, we determined dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) content in retina and vitreous and measured mRNA expression levels of selected clock and circadian rhythm-related genes in the retina/RPE. Results Myopia developed in the goggled eyes of all cohorts. Whereas outdoor rearing lessened myopia by 44% at 4 days, a protective effect was no longer evident at 11 days. Outdoor rearing had no consistent effect on retinal or vitreous content of dopamine or DOPAC. Conforming to prior data on form-deprivation myopia, retina and vitreous levels of DOPAC were reduced in goggled eyes. Compared with contralateral eyes, the retinal expression of clock and circadian rhythm-related genes was modestly altered in myopic eyes of chicks reared indoors or outdoors. Conclusions Outdoor rearing of chicks induces only a partial decrease of goggle-induced myopia that is not maintained, without evidence that retinal dopamine metabolism accounts for the partial myopia inhibition under these outdoor conditions. Although modest, alterations in retinal gene expression suggest that studying circadian signals might be informative for understanding refractive mechanisms.

An immunohistochemically distinct population of cat ciliary ganglion cells

In the cat ciliary ganglion, 16% and 23% respectively of the neurons are surrounded by nerve fibers immunoreactive to calcitonin gene-related peptide (CGRP) and somatostatin (SOM). One-third of the ganglion cell perikarya are immunoreactive to neuropeptide Y (NPY). Analysis of the coincidence of immunoreactivities shows a striking correlation. Practically all of the ganglion cells surrounded by CGRP-like immunoreactive (LI) nerve fibers also are surrounded by SOM-LI nerve processes and demonstrate NPY-LI perikarya. These observations define a subset of NPY-LI ciliary ganglion cells with a particular peptidergic input and perhaps a specific physiologic function.

Substance P-like Immunoreactive Nerves in the Anterior Segment of the Eye

The cholinergic and adrenergic components of the autonomic nervous system regulate important physiologic processes in the anterior segment of the eye. Not only do they control pupil size [l]; they also appear to influence blood flow [2], aqueous humor secretion [3] and aqueous humor outflow [4, 5].