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Intracellular calcium concentration ($[Ca^{2+}]_i$) may play a crucial role in a variety of neuronal functions. Here we report that in primary culture of mouse cerebellar granule cells nicotinic acetylcholine receptors (nAChRs) are expressed in a specific developmental stage and involved ...

Intracellular calcium concentration ($[Ca^{2+}]_i$) may play a crucial role in a variety of neuronal functions. Here we report that in primary culture of mouse cerebellar granule cells nicotinic acetylcholine receptors (nAChRs) are expressed in a specific developmental stage and involved in the regulation of intracellular calcium homeostasis. Nicotine-mediated calcium responses were measured using $^{45}Ca^{2+}$ or fluorometrically using the calcium-sensitive fluorescent dye fura-2. Maximal uptake of $^{45}Ca^{2+}$ evoked by nicotine in mouse cerebellar granule cells were revealed $8{\sim}12$ days in culture. In contrast, nicotine did not alter the basal $^{45}Ca^{2+}$ uptake in cultured glial cells. In cerebellar granule cells nicotine-evoked $^{45}Ca^{2+}$ uptake was largely blocked by the NMDA receptor antagonists. Glutamate pyruvate transaminase (GPT). which removes endogenous glutamate, also prevented nicotine effects, implying the indirect involvement of glutamate in nicotine-mediated calcium responses. Fluorometric studies using fura-2 showed two phases of nicotine-evoked $[Ca^{2+}]_i$ rises: the initial rising phase and the later plateau phase. Interestingly, the NMDA receptor antagonists and GPT appeared to inhibit only the later plateau phase of nicotine-evoked $[Ca^{2+}]_i$ rises. The present results imply that nicotine mediated $^{45}Ca^{2+}$ uptake and $[Ca^{2+}]_i$ rises are attributed to the calcium fluxes through both nAchRs and NMDA receptors in a time-dependent manner. Consequently, nAChRs may play an important role in neuronal development by being expressed in a specific developmental stage and regulating the intracellular calcium homeostasis.

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