Synaptic transmission between rat spinal cord explants and dissociated superior cervical ganglion neurons in tissue culture

Abstract

Physiological properties of the synapses formed between explants of spinal cord and dissociated autonomic ganglion neurons in tissue culture were studied using intracellular and extracellular stimulation and recording techniques (as well as iontophoresis) with a culture perfusion system allowing continuous microscopic observation during repeated changes of the bathing medium. The principal neurons of the superior cervical ganglion (SCGN) were dissociated from perinatal rats and the spinal cord explants were obtained from 15-day rat fetuses; these were allowed to mature for 3-10 weeks in co-culture. Recordings from over 1000 SCGN established that: (a) spontaneous small depolarizations and action potentials occurred in 20% of the SCGN studied, (b) the EPSPs observed in SCGN after spinal cord stimulation were sensitive to decreased Ca2+ and increased Mg2+, as well as to D-tubocurare, hexamethonium and mecamylamine, but not to atropine (at 10(-6) M concentration) or to the alpha-adrenergic blocking agents phentolamine or phenoxybenzamine; no potentiation of the EPSPs was seen with neostigmate or eserine, (c) acetylcholine directly applied to the SCGN was seen to mimic the responses seen after spinal cord stimulation; tetrodotoxin blocked both direct and iontophoretically fired action potentials, with only a suprathreshold acetylcholine potential remaining. These synapses were not sensitive to alpha-bungarotoxin. It is concluded that the synapses formed by spinal cord neurites on principal SCGN in tissue culture are nicotinic cholinergic, and that the evoked EPSPs recorded in this study are thus similar to the orthodromic fast EPSPs observed in vivo. No slow synaptic responses were observed and no demonstrable effects were noted that could be attributed to adrenergic transmission.