Capsaicin is a pungent pain-producing compound found in plants of the capsicum family; it exerts excitatory, desensitizing, and toxic effects on a subset of sensory neurons, including the polymodal nociceptor population. We have carried out a quantitative study of capsaicin-induced fluxes of sodium, guanidine, calcium, rubidium, and chloride ions in cultures of neonatal and adult rat DRG neurons, in conjunction with the use of a histochemical stain that identifies capsaicin-sensitive neurons by means of cobalt uptake. Those cells that take up cobalt in a capsaicin-dependent manner (EC50 = 0.2 microM) represent about 50% of the total neuronal population derived from neonatal DRGs on short-term culture. Overnight treatment of cultures with 2 microM capsaicin leads to the loss of the cobalt-staining subpopulation. The capsaicin-insensitive neurons contain immunoreactive neurofilament epitopes that are present in fewer than 10% of capsaicin-sensitive neurons. This observation provides indirect evidence that the sensitive cells correspond to the small, dark B-type neurons, which are negative for neurofilament immunoreactivity in vivo. A capsaicin-dependent calcium uptake (EC50 = 0.2 microM), as measured by 45Ca incorporation, is shown by a DRG neuronal subpopulation that, like the cobalt-staining population of DRG neurons, is lost after overnight capsaicin treatment (2 microM). Capsaicin application leads to the accumulation of millimolar levels of calcium within a few minutes. Cadmium and other divalent cations block capsaicin-induced calcium uptake, but little or no inhibition is seen with organic calcium channel antagonists. Mitochondria, rather than the endoplasmic reticulum, are the probable destination of the internalized calcium, because ruthenium red inhibits calcium uptake (IC50 = 0.05 microM), whereas methylxanthines are inactive. The subset of sensory neurons that takes up calcium also releases 86Rb when exposed to capsaicin (EC50 = 0.06 microM). No efflux of 36Cl ions could be induced by capsaicin. These cells also show a capsaicin-induced uptake of 22Na or 14C guanidine (EC50 = 0.06 microM). In contrast, chick DRG cells in culture showed no capsaicin-induced calcium or cobalt uptake. Primary cultures of rat superior cervical ganglion neurons and Schwann cells, and a number of neuronal cell lines, also failed to respond to capsaicin, as judged by the calcium, cobalt, or guanidine uptake assays.

SUMMARY1. The responses of adult and neonatal rat dorsal root ganglion (DRG) neurones to buffered acidic solutions were studied with both voltage clamp and radioactive ion flux techniques. Electrophysiological experiments were made on acutely isolated neurones and ion flux experiments were made on cells that had been in culture for 3-6 days.2. Acid solutions of pH < 6-2 evoked a sustained, slowly inactivating inward current in neurones voltage clamped at negative holding potentials. The size of the current increased with increasing proton concentrations. This response was restricted to a sub-population (-45°%) of adult and neonatal rat DRG neurones and was distinct from a rapidly activating and inactivating proton-induced inward sodium current that was also found in DRG neurones.3. The proton-activated sustained current was due to an increase in cation conductance that allowed K+, Cs' and Na+ to pass with PK/PNa = 1-32 and Pcs/PNa = 142.

1 We have found that capsazepine, a competitive antagonist at the vanilloid (capsaicin) receptor, blocks voltage-activated calcium currents in sensory neurones. 2 The block of calcium current was slow to develop with a half time of about one minute at 100 mM and lasted for the duration of the experiment. The rate of block of calcium current was strongly concentration-dependent. 3 The EC 50 for the blocking e ect at 0 mV was 7.7+1.4 mM after 6 min exposure to capsazepine. The EC 50 at equilibrium was estimated to be 1.4+0.2 mM. 4 The block of calcium current showed some voltage-dependence but there was no indication of any selectivity of action for a calcium channel subtype. The characteristics of the blocking action of capsazepine on the residual current of cells which were pretreated with either o-conotoxin or nimodipine were similar to control. 5 The data suggest that capsazepine, in addition to its competitive antagonism of vanilloid receptors, has a non-speci®c blocking action on voltage-activated calcium channels which should be taken into account when interpreting the e ects of this substance on intact preparations in vitro or in vivo.

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