Excitatory amino acid transporter 5 is widely expressed in peripheral tissues.

It is routinely stated in the literature that Excitatory Amino Acid Transporter 5 (EAAT5) is a retina-specific glutamate transporter. EAAT5 is expressed by retinal photoreceptors and bipolar cells, where it serves as a slow transporter and as an inhibitory glutamate receptor, the latter role is due to the gating of a large chloride conductance. The dogma of an exclusively retinal distribution has arisen because Northern blot analyses have previously shown only modest hybridisation in non-retinal tissues. Others have re-interpreted this as indicating that EAAT5 was only present in retinal tissues. However, this view appears to be erroneous; recent evidence demonstrating abundant expression of EAAT5 in rat testis prompted us to re-examine this dogma. A new antibody was developed to an intracellular loop region of rat EAAT5. This new tool, in concert with RT-PCR and sequencing, demonstrated that EAAT5 is widely distributed at the mRNA and protein levels in many non-nervous tissues including liver, kidney, intestine, heart, lung, and skeletal muscle. We conclude that EAAT5 is a widely distributed protein. Whether it functions in all locations as a glutamate transporter, or mainly as a glutamate-gated chloride conductance, remains to be determined.

Selective inner retinal dysfunction precedes ganglion cell loss in a mouse glaucoma model.

BACKGROUND/AIMS: To correlate ganglion cell function with defined parameters of the elevated intraocular pressure profile (IOP) in a mouse glaucoma model and to determine the temporal relationship of these functional changes with ganglion cell death. METHODS: Unilateral chronic ocular hypertension was induced in C57BL6/J mice by laser ablation of the limbal episcleral veins. Scotopic flash electroretinograms were recorded after 5, 10, 20, and 40 days to isolate specific outer and inner retinal responses. Inner retinal function was correlated with the pressure differential between treated and non-treated eyes at the time of electroretinographic recording, and with the cumulative IOP insult (the integral of the IOP.time profile). Peripheral and central ganglion cell densities were quantified by Brn-3 immunohistochemistry. RESULTS: Elevated IOP induced a preferential deficit in inner retinal function. The positive scotopic threshold response (pSTR) was suppressed by 68% on day 5, by 50% on day 10, by 54% on day 20 and by 46% on day 40 after laser treatment. Inhibition of the STR correlated with the pressure differential between treated and non-treated eyes but not with the IOP.time integral. Inner retinal dysfunction preceded the progressive death of ganglion cells. Ganglion cell loss occurred preferentially in peripheral retina and correlated with the cumulative IOP insult. CONCLUSION: We have demonstrated specific inner retinal dysfunction in an inducible mouse glaucoma model. STRs are sensitive to elevated IOP per se, and their early suppression reflects ganglion cell dysfunction rather than cell death. The correlation between IOP elevation and suppression of inner retinal function, in the context of the temporal progression of ganglion cell death, suggests that a portion of the IOP-mediated ganglion cell dysfunction may be reversible.

A neurological symptom survey of patients with type I Gaucher disease.

Gaucher disease is an inborn error of glycosphingolipid metabolism resulting from deficiency of the lysosomal enzyme glucocerebrosidase. The majority of the patients (with type I disease) do not have primary central nervous system involvement. However, several studies have noted that secondary neurological complications may develop as a consequence of nerve root or spinal cord compression following vertebral body collapse or, for those with coagulation disorders, bleeding within confined compartments. An epidemiological survey was conducted to ascertain the incidence of neurological symptoms in patients with Gaucher disease type I (GD I). The survey included a review of the patients' medical history, an estimate of Gaucher disease severity according to a modified Symptom Severity Index (SSI), and completion of a questionnaire regarding their neurological status and Quality of Life (QoL) according to the SF-36 Health Survey. Seventy-three per cent of respondents were found to have experienced at least one neurological complaint in the preceding 3 months. Adult patients with Gaucher disease often have other medical problems unrelated to their primary diagnosis. Thus, the high incidence of neurological complaints in these patients may be attributable to concurrent medical problems and/or side-effects from concomitant medications. These issues may influence patients' assessment of their disease severity and/or response to treatment.

Differential perturbation of neuronal and glial glutamate transport systems in retinal ischaemia.

Glutamate is the major excitatory neurotransmitter in the retina and is removed from the extracellular space by an energy-dependent process involving neuronal and glial cell transporters. The radial glial Müller cells express the glutamate transporter, GLAST, and preferentially accumulate glutamate. However, during an ischaemic episode, extracellular glutamate concentrations may rise to excitotoxic levels. Is this catastrophic rise in extracellular glutamate due to a failure of GLAST? Using immunocytochemistry, we monitored the transport of the glutamate transporter substrate, D-aspartate, in the retina under normal and ischaemic conditions. Two models of compromised retinal perfusion were compared: (1) Anaesthetised rats had their carotid arteries occluded for 7 days to produce a chronic reduction in retinal blood flow. Retinal function was assessed by electroretinography. D-aspartate was injected into the eye for 45 min. Following euthanasia, the retina was processed for D-aspartate, GLAST and glutamate immunocytochemistry. Although reduced retinal perfusion suppresses the electroretinogram b-wave, neither retinal histology, GLAST expression, nor the ability of Müller cells to uptake D-aspartate is affected. As this insult does not appear to cause excitotoxic neuronal damage, these data suggest that GLAST function and glutamate clearance are maintained during periods of reduced retinal perfusion. (2) Occlusion of the central retinal artery for 60 min abolishes retinal perfusion, inducing histological damage and electroretinogram suppression. Although GLAST expression appears to be normal, its ability to transport D-aspartate into Müller cells is greatly reduced. Interestingly, D-aspartate is transported into neuronal cells, i.e. photoreceptors, bipolar and ganglion cells. This suggests that while GLAST is vitally important for the clearance of excess extracellular glutamate, its capability to sustain inward transport is particularly susceptible to an acute ischaemic attack. Manipulation of GLAST function could alleviate the degeneration and blindness that result from ischaemic retinal disease.

Combined effect of cyclosporine and sirolimus on improving the longevity of recombinant adenovirus-mediated transgene expression in the retina.

OBJECTIVES: To reevaluate the longevity and intraocular safety of recombinant adenovirus (rAd)-mediated gene delivery after subretinal injection, and to prolong transgene expression through the combination of 2 synergistic immunosuppressants. METHODS: An rAd vector carrying green fluorescent protein (GFP) gene was delivered subretinally in the rat eye. The GFP expression was monitored in real time by fundus fluorescent photography. Intraocular safety was examined by observation of changes of retinal pigmentation, cell infiltration in virus-contacted area, immunophenotyping for CD4(+) and CD8(+) cytotoxic T lymphocytes, and CD68(+) macrophages, histologic findings, and dark-adapted electroretinography. Two synergistic immunosuppressants, cyclosporine and sirolimus, were used alone or in combination to prolong transgene expression by temporary immunosuppression. RESULTS: The GFP expression peaked on day 4, dramatically decreased on day 10, and was not detectable on day 14. The decreased GFP expression was coincident with cell infiltration in virus-contacted area. Immunostaining showed that the infiltrating cells were CD4(+) and CD8(+) cytotoxic T lymphocytes and CD68(+) macrophages. Clumped retinal pigmentation and decreased b wave of dark-adapted electroretinogram were observed at 3 to 4 weeks after injection. Histologic examination confirmed rAd-induced retinal degeneration. Transient immunosuppression by cyclosporine and sirolimus, either alone or in combination, improved transgene expression, with the combination being the most efficient. The combined immunosuppression attenuated but did not retard the rAd-induced retinal damage. CONCLUSIONS: Transgene expression mediated by rAd after subretinal delivery is short-term and toxic to the retina. Combination of cyclosporine and sirolimus may act as an immunosuppressive adjunct to prolong rAd-mediated gene transfer. CLINICAL RELEVANCE: The intraocular safety of rAd should be carefully considered before clinical trials are performed.

Expression of the beta6 integrin subunit is associated with sites of neutrophil influx in lung epithelium.

Inhalation of ozone by Rhesus monkeys results in epithelial injury and granulocyte influx in both conducting airways and respiratory bronchioles. We have reported that ozone-induced neutrophil recruitment and subsequent epithelial repair can be inhibited in vivo with a CD18 antibody. The antibody-mediated effect is abrogated by local instillation of C5a (a CD18-independent neutrophil chemoattractant), thereby demonstrating a role for neutrophils in lung epithelial repair processes. As an extension of this study, we examined the effect of ozone and neutrophil influx on epithelial expression of the beta6 integrin, an adhesion molecule associated with proliferation and repair. Expression of beta6 integrin was determined by immunohistochemistry for ozone-exposed monkeys treated with either control immunoglobulins or a CD18 antibody. The tracheal epithelium of ozone-exposed monkeys treated with control immunglobulins expressed the beta6 integrin. In contrast, the tracheal epithelium of ozone-exposed monkeys treated with CD18 antibody exhibited very low to undetectable expression of beta6 integrin. In association with C5a instillation and neutrophil influx, beta6 integrin was also observed in respiratory bronchiolar epithelium from both control and ozone-exposed animals. These findings cumulatively suggest that lung epithelial cell expression of beta6 integrin is associated with sites of neutrophil recruitment.

Long-term real-time monitoring of adeno-associated virus-mediated gene expression in the rat retina.

PURPOSE: Previous studies have demonstrated that adeno-associated virus (AAV) efficiently transduced retinal pigmented epithelial (RPE) cells. The goal of this study was to further evaluate and characterize transgene expression within the RPE cells over time in vivo. METHODS: Adeno-associated virus-mediated gene transfer was monitored and quantified by retinal photography following subretinal injection of a recombinant AAV encoding the green fluorescent protein gene (rAAVCMV-gfp) into rat eyes. Retinal function of transduced rat eyes was measured by electroretinography. RESULTS: The maximum level of transgene expression was reached at 8 weeks postinjection followed by a gradual decrease throughout the experimental period. Interestingly, it was observed that while gfp expression was stable in some RPE cells, gfp fluorescence completely disappeared in other cells over the duration of the experiment. The expression of AAV-mediated gfp in RPE cells did not alter the retinal function for over 1 year CONCLUSIONS: These results confirm the importance of this direct visualization system to study vector transgene expression in vivo and support the use of AAV for diseases treatable by targeting RPE cells.

Over-the-counter medicines and the elderly.

Sales of over-the-counter (OTC) medicines are rising, and will continue to rise as more products are reclassified from prescription-only status to OTC medicines (either pharmacy-only or general sales list). Patients and doctors often omit discussion of OTC medicines when giving or taking a medication history. This has serious potential for identifying adverse drug reactions and drug-drug interactions, which are more common in older people. Therefore, medication histories should include documentation of any OTC medicines taken.

Are neuronal transporters relevant in retinal glutamate homeostasis?

Exposure of isolated retinas to 30 microM D-aspartate, which is a substrate for all high affinity glutamate transporters, for 30 min, resulted in the accumulation of such D-aspartate into Müller glial cells but not glutamatergic neurons as evinced by immunocytochemistry for D-aspartate. Further incubation of such loaded retinas in physiological media, in the absence of D-aspartate, resulted in the slow release of accumulated D-aspartate from the Müller cells and its accumulation into populations of photoreceptors and bipolar cells. This result indicates that after initial transport into Müller cells, reversal of direction of transport of D-aspartate, and thus by inference glutamate, by GLAST, readily occurs. D-aspartate released by Müller cells was strongly accumulated into cone photoreceptors which are known to express GLT-1, and into rod photoreceptors which we demonstrate here to express the retina specific glutamate transporter EAAT5 (excitatory amino transporter 5). Populations of glutamatergic bipolar cells, which express GLT-1 also exhibited avid uptake of D-aspartate. We conclude that the Müller cell glutamate transporter GLAST is responsible for most of the initial glutamate clearance in the retina after its release from neurones. However, some glutamate is also returned from Müller cells, to neurons expressing GLT-1 and EAAT5, albeit at a slow rate. These data suggest that the role of neuronal glutamate transporters in the retina may be to facilitate a slow process of recycling glutamate back from Müller cells to neurons after its initial clearance from perisynaptic regions by GLAST.

Inhibition of Müller cell glutamine synthetase rapidly impairs the retinal response to light.

It is widely assumed that neurones have sufficient metabolic reserves to allow them to function independently of glial cells for extended periods. The present study investigates the length of time taken before retinal neurones no longer respond normally to light after the inhibition of glial enzymes that are involved in the synthesis of precursors of neuronal glutamate. The glutamine synthetase inhibitor methionine sulfoximine, when injected intraocularly in Wistar rats, caused a time- and dose-dependent suppression of the scotopic electroretinogram b-wave. At the highest dosage (40 mM) the b-wave was significantly reduced within 2 min of injection. Because the b-wave is an indicator of neurotransmission in the retina, it is deduced that inhibition of glutamine synthetase rapidly blocks glutamatergic neurotransmission. Immunohistochemistry revealed a depletion of neuronal glutamate and an accumulation of glutamate in Müller glial cells, in a time course that matched the b-wave suppression. The b-wave was quickly restored by injection of glutamine (4 mM). The rapid reduction of glutamatergic transmission after methionine sulfoximine administration challenges the view that neurones have sufficient reserves to allow them to function independently for extended periods; instead, it indicates that glia are essential for the moment-to-moment sustenance of neuronal function.

Developmental expression of excitatory amino acid transporter 5: a photoreceptor and bipolar cell glutamate transporter in rat retina.

Excitatory amino acid transporter 5 (EAAT5) is a retina-specific glutamate transporter which has an associated chloride conductance. Thus it is comparable in its functional properties to the glutamate transport systems previously described in photoreceptors and some bipolar cells. We have raised antibodies to the carboxyl- and amino-terminal regions of EAAT5. Labeling for both of these antisera was developmentally regulated: weak labeling appeared in photoreceptors around P7; by P10 strong labeling was present in photoreceptors and by P21 a population of bipolar elements were also weakly labeled. In adult retinae both antisera heavily immunolabeled all photoreceptors as well as a heterogeneous population of bipolar cell somata and their proximal axonal processes: synaptic terminals of these cells were also labeled after partial proteolytic digestion of the tissues. The positions and morphology of these terminals suggests that they are the terminals of both rod and cone rod bipolar cells. We conclude that in rat retina, EAAT5 is a photoreceptor and bipolar cell glutamate transporter.

Antisense knockdown of GLAST, a glial glutamate transporter, compromises retinal function.

PURPOSE: To elucidate the role of the glial glutamate transporter GLAST, in the regulation of retinal function. METHODS: Antisense oligonucleotides to GLAST were injected intravitreally into the left eye of Wistar rats. Sense oligonucleotides (control) were injected into the right eye over a period of 3 days. Scotopic flash electroretinograms were recorded over a 20-day period. To assay whether the antisense oligonucleotides caused a reduction in the expression or the activity of GLAST, retinas were exposed to D-aspartate, a nonendogenous substrate of glutamate transporters. The retinas were immunolabeled with specific antibodies for D-aspartate. Retinal GLAST and glutamate distributions also were determined immunocytochemically. RESULTS: Antisense oligonucleotides markedly suppressed the electroretinogram b-wave, whereas sense oligonucleotides had no significant effect. Significant changes in the electroretinogram were apparent 5 days after injection of antisense oligonucleotide and were sustained for at least 20 days. A marked reduction of D-aspartate uptake into Muller cells of retinas that had been exposed to the antisense oligonucleotides 5 days previously suggests a reduction of GLAST activity. The retinas, however, displayed no evidence of excitotoxic neuronal degeneration, and the distribution of glutamate was unaffected by antisense treatment. CONCLUSIONS: The observed lack of neuronal degeneration suggests that reduced glutamate uptake into Muller cells does not cause excitotoxic tissue damage. A direct perturbation of glutamatergic signaling is more likely, because the rapid clearance of glutamate is necessary for light elicited signaling between photoreceptors and bipolar cells. This suggests that GLAST is essential for the maintenance of normal retinal transmission.

Changing patterns of spatial buffering of glutamate in developing rat retinae are mediated by the Müller cell glutamate transporter GLAST.

The patterns of expression of the glutamate transporter GLAST were compared with the patterns of uptake of exogenous D-aspartate, which is a substrate for all glutamate transporters. At postnatal day 0, fine radial processes and end feet of presumptive Müller cells were weakly immunoreactive for GLAST. At postnatal day 3, intense labelling was associated with astrocytes enveloping newly formed blood vessels on the vitread surface of the retina. Between postnatal days 7 and 10, there was a rapid increase in the intensity of labelling in the Müller cells but clear stratification of GLAST-immunoreactive processes in the inner plexiform layer was not observed until postnatal day 14. By comparison, D-aspartate uptake was initially associated with a wide variety of cellular elements including most neuroblasts, presumptive Müller cells, and astrocytes associated with blood vessels but was absent from the somata of many neurons in the ganglion cell layer and amacrine cell layer. There was a gradual contraction in the numbers of cells that were able to take up D-aspartate, such that, by adulthood, uptake was restricted mainly to Müller cells and astrocytes. We conclude that, during early retinal development, the low levels of GLAST expression by Müller cells permit D-aspartate, and by inference, glutamate, to permeate the retina freely, thus allowing uptake by other glutamate transporters on other cell types. As the retina matures, increased expression of GLAST by Müller cells restricts the access of D-aspartate to other cellular compartments in the retina. This changing pattern of spatial buffering of glutamate by GLAST probably has significant implications regarding our understanding of the role of glutamate during processes such as retinal synaptogenesis.

Drug therapy and the older person: role of the pharmacist.

Older people in the UK receive a disproportionate amount of medication. They comprise 18% of the population but receive 45% of all prescription items. Not surprisingly they experience drug-related illnesses - in 1980, 1 in 10 admissions to acute geriatric units were wholly or partly due to adverse drug reactions. Drugs which should be used with particular care or even avoided in older people include benzodiazepines, warfarin, digoxin, aminoglycosides, tricylic antidepressants, antipsychotics and long-acting oral hypoglycaemic agents. Pharmacists can promote safer prescribing practices by advising both patients and doctors. The community pharmacist can assist in drug compliance by providing patients with additional information about individual drugs, identifying potential adverse drug reactions and interactions, supplying appropriate drug containers or compliance aids, and even arranging home visits for patients unable to visit the pharmacist. Some community pharmacists provide pharmaceutical advice and services to residential and nursing homes. Pharmacists' advice to doctors can include one to one discussions in either primary or secondary care, assisting in medication review, providing information to prescribing committees, compiling drug formularies, assisting in auditing of prescribing practices and organising disposal of unwanted medicines and poisons campaigns.

Chrysiasis revisited: a clinical and pathological study.

Chrysiasis is a distinctive and permanent pigmentation of light-exposed skin resulting from the administration of parenteral gold salts. We report a study of 40 Caucasian patients with rheumatoid arthritis, treated with intramuscular sodium aurothiomalate, of whom 31 had chrysiasis. Visible changes develop above a threshold, equivalent to 20 mg/kg gold content, and their severity depends upon cumulative dose. Focal aggregates of particulate gold are deposited in the reticular and papillary dermis in amounts that correlate with the degree of pigmentation. Characteristically, initially the periorbital region is affected by a mauve discoloration, which intensifies and deepens into a blue/slate-grey colour, while extending to involve the face, neck and upper limbs. Although chrysiasis develops insidiously and patients may be unaware of the changes, positive identification is important in order to avoid misdiagnosis and medical mismanagement, and afford appropriate reassurance. Prevention is difficult, but measures to reduce sunlight exposure may be helpful.

Influence of excitatory amino acids and ischemia on rat retinal choline acetyltransferase-containing cells.

PURPOSE: To compare the effects of glutamate agonists and different types of ischemic insult on choline acetyltransferase (ChAT) immunoreactivity in the rat retina. METHODS: Rat retinas were exposed to different glutamate agonists in vivo or in vitro for specific periods of time, and the retinas were then fixed and processed for the localization of ChAT immunoreactivity. In other experiments, rats were administered an ischemic insult either by ligaturing the carotids (two-vessel occlusion [2-VO] procedure), cannulating the anterior chamber, and raising the intraocular pressure (high intraocular pressure [HIOP] procedure) or placing a ligature around the optic nerve sufficiently tightly to prevent blood flow through the central retinal artery (vascular ligation [VL] procedure). The electroretinogram was recorded, and, after a specific period of time, reperfusion was allowed to occur. Thirty to 36 hours after reperfusion, the retinas were dissected and processed for the localization of ChAT, as well as for parvalbumin, Thy-1, and alpha PKC immunoreactivities. RESULTS: Of the glutamate agonists tested, only kainate reduced ChAT immunoreactivity significantly in vivo and in vitro. This effect of kainate could be counteracted by the antagonist CNQX (6-cyano-2,3-dihydroxy-7-nitroquinoxaline-2,3-dione). The ChAT immunoreactivity was unaffected in retinas in which ischemia was induced by the 2-VO procedure. In contrast, ChAT immunoreactivity was obliterated in retinas in which the HIOP was used and drastically reduced when the VL procedure was used. Interestingly, neither alpha PKC nor Thy-1 immunoreactivities were affected in retinas subjected to HIOP or VL methods. However, parvalbumin immunoreactivity was reduced in the HIOP model but only slightly altered in the VL model. CONCLUSIONS: The current results suggest that kainate receptors are associated with the cholinergic retinal neurones in the rat retina. Activation of these receptors by kainate causes a reduction in the neurones' ChAT content. This effect can be mimicked by subjecting the retina to a sufficiently harsh ischemic insult, as occurs in the VL and HIOP procedures. When the ischemic insult is mild, as in the 2-VO procedure, no obvious change in ChAT immunoreactivity is apparent. The HIOP procedure for inducing an ischemic insult was found to be the most severe of the three procedures used, because ChAT immunoreactivity was obliterated and clear changes in the parvalbumin immunoreactivity also were recorded. Interestingly, neither the HIOP nor the VL procedures caused a change in the Thy-1 and alpha PKC immunoreactivities.

Prolonged bilateral carotid artery occlusion induces electrophysiological and immunohistochemical changes to the rat retina without causing histological damage.

Reduction of the retinal blood flow by occlusion of both common carotid arteries suppressed the b-wave of the rat's electroretinogram. Transient occlusion of the carotids for 45 min reduced the b-wave by 46% without affecting the amplitude of the a-wave. The normal ERG activity returned 30 min after restoration of blood flow. Prolonged carotid occlusion for 7 days totally abolished the b-wave but enhanced the a-wave amplitude. Although b-wave amplitude suppression has been considered as an indicator of retinal ischaemia, no histological changes were seen in retinas of rats subjected to 45 min or 7 days of two-vessel occlusion, when observed by light microscopy. Moreover, GABA, GABAA receptor, calretinin and PKC-alpha immunoreactivities were unaltered. Carotid artery occlusion did, however, induce the expression of the cytoskeletal protein, glial fibrillary acidic protein (GFAP), in retinal Müller cells. The increase in the Müller cell GFAP immunoreactivity was related to how long the carotids were occluded as well as the reperfusion time. Prolonged occlusion for 7 days resulted in a 356% increase in retinal GFAP. These findings show that a reduction of retinal blood flow by occlusion of the carotids causes a metabolic stress to the retina and elicits events associated with gliosis without resulting in 'ischaemic-like' morphological changes.

Redistribution of GABA immunoreactivity following central retinal artery occlusion.

gamma-Aminobutyric acid (GABA) is normally primarily in amacrine cells in the rat retina. Immediately after an ischaemic insult, attained by occlusion of the central retinal artery for 60 min, GABA is then found to be associated with Müller cells. During subsequent reperfusion, the distribution of GABA immunoreactivity gradually reverts from the glial cells back into neuronal elements of the retina. Twenty-four hours after ischaemia, GABA staining is indistinguishable from that seen in control animals. It is suggested that during central retinal artery occlusion, Müller cell energy levels are sufficient to allow the active uptake of released GABA, but insufficient to metabolise it to glutamine. The normal cycle of GABA metabolites from Müller cells to neurones is thus inhibited. Restoration of blood flow and the consequent increase in retinal energy levels, as indicated by a slight recovery of the electroretinogram b-wave, facilitates glutamine shunting between glial cells and amacrine cells, resulting in the synthesis of neuronal GABA.

The presence of serotonin (5-HT1) receptors negatively coupled to adenylate cyclase in rabbit and human iris-ciliary processes.

Serotonin (5-hydroxytryptamine, 5-HT) reduces forskolin-induced stimulation of cyclic AMP in rabbit iris-ciliary body (ICB) homogenates. The effect is dose dependent and can be mimicked by a number of 5-HT1 receptor agonists including 5-carboxamidotryptamine (5-CT) and RU 24969 [5-methoxy-3-(1,2,3,6, tetrahydro-4-pyridinyl)-1-indole]. The inhibitory effects of 5-CT and the 5-HT1A selective agent 8-hydroxy-2-(di-n-propyl-amino) tetralin (8-OH-DPAT) on forskolin stimulated adenylate cyclase activity are greater in isolated ciliary processes than in the iris musculature. Spiperone and propranolol significantly antagonize the action of 5-CT in the iris-ciliary body, while ketanserin (5-HT2 antagonist) and ICS 205930 (5-HT3/4 blocker) were without influence, indicating the presence of the 5-HT1A subtype of receptor. Studies carried out on human ICB homogenates also suggest the presence of 5-HT1A-like receptors, although these receptors are not identical to those in rabbit. Similarities include dose-dependent decreases in cAMP levels stimulated by forskolin elicited by 1-(3-chlorophenyl) piperazine (mCPP), 5-CT and 8-OH-DPAT. Moreover, the inhibitory effect of 5-CT can also be significantly reduced by the 5-HT1 receptor antagonist, propranolol. However, unlike the case of rabbit tissue, spiperone was ineffective in abolishing the 5-CT response in human ICB homogenates.

The effect of serotonin on the rabbit isolated iris sphincter muscle.

The rabbit isolated iris sphincter muscle maintained in an isotonic state is unaffected by applied serotonin (5-hydroxytryptamine or 5-HT) whereas carbachol causes the muscle to contract. Serotonin does, however, produce a relaxation of the contracted muscle in a dose-dependent manner. This effect is also induced by the 5-HT receptor agonists 8-OH-DPAT (8-hydroxy-2-[di-n-propyl-amino] tetralin, RU 24969 (5-methoxy-3-[1,2,3,6, tetrahydro-4-pyridinyl]-1-indole) and ipsapirone, suggesting the involvement of 5-HT1A receptors. This view is supported by the finding that metergoline, methysergide and propranolol all counteracted the effect produced by serotonin. While 5-HT3 receptors are not involved in the described process, a minor involvement of 5-HT2 receptors cannot be excluded as methysergide partially counteracted the serotonin response. These data provide evidence that serotonin receptors, in particular the 5-HT1A subtype, may be associated with the iris sphincter muscle and suggest their involvement in the regulation of pupil size.