Clathrin light chain and synaptotagmin I in rat sympathetic neurons

Abstract

Clathrin light chain (clathrin LC) and synaptotagmin I in sympathetic neurons in rat superior cervical ganglia (SCG) were studied using immunofluorescence and confocal microscopy. The distributions of clathrin LC and synaptotagmin I were compared with that of tyrosine hydroxylase (TH) and neuropeptide Y (NPY) in double label experiments. The influence of preganglionic regulation on the expression of clathrin LC and synaptotagmin I in post-ganglionic adrenergic neurons was investigated after cutting the cervical sympathetic trunk. In SCGs and irides of control animals, the clathrin LC- and synaptotagmin-I-positive structures were present in a granular pattern in nerve fibers and varicose terminals. In principal neurons, the two proteins were present in a perinuclear network (the Golgi complex). After decentralization, the synaptotagmin-I- and clathrin LC-positive granules normally present in preganglionic nerve terminals outlining the neuronal somata were no longer observed on day 1, but reappeared, and were increased above control in number and intensity, in axon bundles in the ganglia, on day 3 and up to day 28 post-decentralization. In irides, the fluorescence intensity and density of clathrin LC- and synaptotagmin-I-positive nerve terminals in the dilator plate, were semi-quantified using the confocal microscopy software. It was found that both proteins increased shortly after decentralization. Immunoblot data confirmed the immunohistochemical/confocal microscopy observations. Fast axonal transport of clathrin LC- and synaptotagmin I in preganglionic sympathetic neurons was demonstrated in crush-operated cervical sympathetic trunk. Both proteins rapidly accumulated proximally as well as distally to the crush, demonstrating fast anterograde and retrograde axonal transport (recycling). Thus, clathrin LC and synaptotagmin I are normally present in pre- as well as post-ganglionic sympathetic neurons. The colocalization of clathrin LC with synaptotagmin I in the Golgi complex of the adrenergic neurons may imply that clathrin participates in the synthesis/sorting of the fast transported materials in these neurons. Possible explanations for the increase of the two proteins after decentralization are discussed.