EXPRESSION OF SYNAPTIC MEMBRANE-PROTEINS IN GERBIL PINEALOCYTES IN PRIMARY CULTURE

Pinealocytes of various mammalian species contain abundant synaptic-li ke microvesicles (SLMVs) which are considered the endocrine equivalent of neuronal synaptic vesicles, Although the pinealocyte may thus be a suitable cellular model for experimental in vitro studies of SLMVs, n othing is known about the presence of SLMVs in isolated pinealocytes m aintained under tissue culture conditions, In the present investigatio n, we prepared dissociated primary cultures of gerbil pinealocytes to study the expression and distribution of protein components of synapti c vesicles/SLMVs and the presynaptic plasmalemma in pinealocytes kept in vitro, Using immunofluorescence microscopy, we found that cultured pinealocytes readily expressed all synaptic membrane proteins investig ated, i.e., synaptophysin, synaptotagmin I, synaptobrevin II, syntaxin I and SNAP-25, Punctate immunoreactivity for the vesicle-associated p roteins could be detected throughout the cell bodies of pinealocytes a nd was also distributed into all of their processes which began to dev elop within the first days in culture, Outgrowing processes exhibited growth cone-like structures which were enriched in synaptic vesicle-as sociated proteins, After 1 week in vitro, pinealocytes had frequently formed an elaborate network of long interwoven processes, Accumulation s of synaptic vesicle-associated proteins were observed in varicositie s and terminal swellings of the processes, The vesicle-rich process sw ellings often established synaptic-like contacts with somata and proce sses of other pinealocytes, Some of the pinealocyte processes possesse d additional axon-like properties as demonstrated by their lack of imm unoreactivity for the somato-dendritic marker MAP2 and the transferrin receptor, The comparison of the staining patterns for synaptophysin a nd the endocytotic marker transferrin receptor by confocal laser scann ing microscopy revealed a largely differential intracellular distribut ion of the two proteins, This may indicate that a substantial fraction of pinealocyte SLMVs by-passes the early endosomal-related recycling pathway of SLMVs, Herewith, we have shown that isolated gerbil pinealo cytes maintained in primary culture can acquire morphological and neur ochemical traits which closely mimick those observed in vivo, In parti cular, these cultures permit experimental studies of the compartment o f pinealocyte SLMVs which seem to make up a major secretory pathway fo r paracrine intrapineal communication. (C) 1997 Wiley-Liss, Inc.