A novel modification of the stimulated cobalt uptake technique has been used to identify rat dorsal root ganglion cells expressing capsaicin and bradykinin receptors. The technique involves incubating intact dorsal root ganglia in vitro in a modified Krebs solution in which cobalt chloride has been substituted for calcium. Activation of dorsal root ganglion cells by capsaicin or bradykinin in the presence of the cobalt ions results in cobalt influx into the excited cells. Histochemical methods were then used to visualize the intracellular accumulation of cobalt, and labelled cells were counted and characterized. Capsaicin (2 μM) or bradykinin (500 nM) applied for 20 min induced cobalt uptake in 13.8 ± 0.6 and 9.6 ± 0.5% of neuronal profiles in dorsal root ganglia (L4), respectively, a significantly larger number than stained in control ganglia (in the absence of agonists: 1.8 ± 0.7%). The longest diameter of the soma of stained dorsal root ganglion cells following capsaicin and bradykinin perfusion were significantly different from each other and from the non-labelled population (17.5 ± 0.7 and24.5 ± 0.2 μm for capsaicin;23.2 ± 0.9 and25.5 ± 0.4 μm for bradykinin; labelled and non-labelled cells, respectively). The distribution of cell diameters revealed that while capsaicin-sensitive cells were exclusively small-sized, bradykinin-sensitive cells were predominantly small and medium sized. The selective bradykinin-2 receptor antagonist HOE-140 (5.0 μM) blocked the bradykinin-induced staining (2.16 ± 0.02%) but not that of capsaicin. The bradykinin-1 agonist [des-Arg9]-bradykinin did not induce any significant increase in stained cells over the control number (2.2 ± 0.7%), Superfusion of the dorsal root ganglia with 50 mM KC1 resulted in staining in 1.1 ± 0.6% of dorsal root ganglion cells which was not significantly different from the control suggesting that voltage dependent calcium channels do not participate in the cobalt influx. This technique provides a novel way to study the subpopulations of dorsal root ganglion cells activated by sensory neuron excitants.
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