Expression Of Somatostatin Receptor 5 And Modulation Of Somatostatin On Presynaptic Activity In Rat Retinal Amacrine Cells

Somatostatin(SRIF),a neuroactive peptide in the central nervous system(CNS), exerts its actions via five subtypes of specific receptors(sst_s) and can work as a neurotransmitter or neuromodulator.In the retina,SRIF is mainly localized to amacrine cells(ACs).Localization of sst_1-4 in the retina has been immunocytochemically examined in a variety of species,but immunocytochemical data on the expression of sst₅ in the retina are scant.This could be partly because of the rather low sst₅ mRNA levels detected in retinas of several species.In addition, because of lack of specific antagonists(except for sst₂),there are still limited data on the functional roles of sst_s in neurons.The emergence of commercial available specific sst₅ antagonist makes the study of functions of sst₅ in the retina possible.In this work,the localization of sst₅ was studied immunocytochemically in rat retinal ACs.Labeling for sst₅ was diffusely distributed throughout the full thickness of the inner plexiform layer(IPL) and formed two distinct fluorescence bands in the distal part of the IPL.Double labeling experiments showed that sst₅ was expressed in GABAergic ACs.It was further shown that labeling for sst₅ was observed in both dopaminergic and cholinergic ACs,stained by tyrosine hydroxylase(TH) and choline acetyltransferase(ChAT),respectively.The immunostaining appeared mainly on the cell membranes and somatodendritic compartments of these ACs.For the cholinergic ACs,weak sst₅-immunoreactivity was also observed in the processes terminating in the IPL.In contrast,no sst₅-immunoreactivity was found in glycinergic AII ACs, stained by parvalbumin(PV).Furthermore.labeling for SRIF was co-localized with sst₅ in both dopaminergic and cholinergic ACs.These results suggest that sst₅ may serve as an autoreceptor or conventional receptor in retinal ACs.Using whole cell patch-clamp technique,we investigated presynaptic modulation by SRIF in cultured rat retinal GABAergic ACs.Miniature inhibitory postsynaptic currents(mIPSCs) were recorded in cultured rat GABAergic ACs and could be completely inhibited by the GABA_A receptor antagonist bicuculline,indicating that mIPSCs were mediated by postsynaptic GABA_A receptors.1μM SRIF reduced the frequency of mIPSCs,and the effect could be in part blocked by co-application of the sst₅ antagonist BIM 23056 and the sst₂ antagonist CYN-154806.SRIF of 1μM slightly reduced the mean amplitude of mIPSCs,but did not affect the mean rise time and decay time.In addition,the frequency of mIPSCs enhanced by elevating extracellular Ca²⁺ concentration([Ca²⁺]_o) was also reduced by 1μM SRIF,and almost all mIPSC events disappeared after eliminating[Ca²⁺]_o,revealing the[Ca²⁺]_o dependance of mIPSCs.Similarly,200μM CdCl₂ blocked mIPSCs,suggesting that voltage-gated calcium channels(VGCCs) may play an important role in mIPSCs. Indeed,10μM nimodipine,a specific L-type calcium channel blocker,obviously suppressed mIPSCs,suggesting the involvement of L-type calcium channels. Furthermore,forskolin(5μM),an activator of adenylyl cyclase,enhanced the frequency of mIPSCs,while the PKA inhibitor Rp-cAMP reduced the frequency of mIPSCs.In the presence of Rp-cAMP SRIF had no effect on mIPSCs.These results suggest that SRIF inhibits the Ca²⁺ influx through VGCCs by activating specific receptors,and thus reduces the presynaptic release of GABA from rat retinal ACs via the cAMP-PKA pathway.In conclusion,in the present work,the expression and functions of sst₅ in the rat retinal ACs are systematically studied using immunocytochemical and whole cell patch-clamp techniques.Almost all GABAergic ACs express sst₅.SRIF may depress the presynaptic GABA release from GABAergic ACs via sst₅ and/or sst₂.