Stimulation of voltage-dependent Ca channels by NO at rat myenteric neurons

The aim of the present study was to characterize the action of the neurotransmitter, NO, on rat myenteric neurons. A NO donor such as GEA 3162 (10 moll) induced an increase in the intracellular Ca concentration as indicated by an increase in the fura-2 ratio in ganglia loaded with this Ca-sensitive fluorescent dye. The effect of GEA 3162 was strongly reduced in the absence of extracellular Ca suggesting an influx of Ca from the extracellular space evoked by NO. A similar nearly complete inhibition was observed in the presence of Ca channel blockers such as Ni (510 moll) or nifedipine (10 moll). Whole-cell patch-clamp recordings confirmed the activation of voltage-dependent Ca channels, measured as inward current carried by Ba, by the NO donor. The peak Ba-carried inward current increased from –100 ± 19 pA to –185 ± 34 pA in the presence of sodium nitroprusside (10 moll). The consequence was a hyperpolarization of the membrane, which was blocked by intracellular Cs, and thus most probably reflects the activation of Ca-dependent K channels. Furthermore, at least two subtypes of NO-synthases, the NOS-1 (neuronal form) and the NOS-3 (endothelial form), were found as transcripts in mRNA isolated from the rat myenteric ganglia. The expression of these NO-synthases was confirmed immunohistochemically. These observations suggest that NO, released from nitrergic neurons within the enteric nervous system, does not only affect target organs such as smooth muscle cells in the gut, but has in addition profound effects on the enteric neurons themselves, the key players in the regulation of many gastrointestinal functions.