Generation of an unusual depolarizing response in rabbit primary afferent neurones in the absence of divalent cations.

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Abstract
The effects of divalent cations on responses to 5-hydroxytryptamine (5-HT), GABA and 1,1-dimethyl-4-phenyl piperazinium (DMPP) were investigated using a sucrose-gap method to record population responses. In Ca-free medium responses to 5-HT were enhanced, those to DMPP depressed and those to GABA unchanged. In Mg-free medium responses to 5-HT were unchanged, while those to DMPP and GABA were depressed. Removal of both Ca and Mg from the superfusion medium caused a small reduction of GABA responses and a large reduction of DMPP responses. Responses to 5-HT were not only greatly potentiated but were changed in character; the depolarizing phase became sigmoid and the dose dependence between quantity of 5-HT and response magnitude was lost as if 5-HT were triggering an all-or-nothing phenomenon. Dose-response relationships for GABA were normal in the large majority of preparations. In .apprx. 10% of preparations, supramaximal amounts of GABA or DMPP evoked large responses of a similar character to those evoked by 5-HT. The large responses, generated by an unknown mechanism, were termed X responses. Further reduction in tissue divalent cations by EGTA [ethylene glycol-bis-(.beta.-aminoethyl ether) N,N''-tetraacetic acid] (1 mM) caused X responses to be generated spontaneously. Ca, Mg, Mn or Co (1 mM) could suppress X responses. DMPP responses, reduced in Ca/Mg-free medium, were largely restored by 1 mM-Ca. Depression of GABA responses in Ca/Mg-free medium could be entirely attributed to the absence of Mg, Mn being able to substitute for Mg. X responses were generated only after equilibration for 1 h with Ca/Mg-free medium. Attempts to manipulate [Ca]i with dinitrophenol or caffeine did not produce the conditions under which X responses were generated. Intracellular records of responses to 5-HT, GABA or DMPP showed that cells with A fibers responded to GABA but not to 5-HT or DMPP. Fifty-four of 67 cells with C fiber axons (80%) were depolarized by 5-HT, 37 of 49 (76%) by DMPP and 40 of 57 (70%) by GABA. Eighteen of 38 (47%) C cells were depolarized by all 3 agents. Some C cells were very sensitive to 5-HT, .mu.M evoking a substantial response. In most, responses to 10-5 M-5-HT had a slower rate of rise than responses to 10-4 or M-GABA or DMPP, yet lower 5-H concentrations normally elicited X responses in sucrose-gap experiments whereas GABA or DMPP normally did not. 5-HT apparently elicits a large depolarization in a few sensitive cells which in turn excite surrounding cells. K efflux may play a role in this process, since increases in [K]o during superfusion with Ca/Mg-free medium initiated X responses. X responses were dependent on [Na]o but were not blocked by tetrodotoxin [TTX] (10-7-10-6 M). Voltage-sensitive Na channels which are TTX insensitive and whose activation threshold is lowered in the absence of divalent cations may be involved.