Molecular Mechanisms of Action Induced by 5-HT₃ Receptors in a Neuronal Cell Line and by 5-HT₂ Receptors in a Glial Cell Line

Summary

The molecular mechanisms of action of serotonin were investigated in a neuronal cell line expressing 5-HT₃ receptors (neuroblastoma × glioma hybrid cells) and in a glioma cell line with 5-HT₂ receptors. In both cell lines, serotonin induces a transient rise of cytosolic Ca²⁺ activity. Ca²⁺ channel blockers (Ni²⁺ and La²⁺) suppress the Ca²⁺ response to serotonin in the neuronal cells but not in the glial cells. When internal Ca²⁺ stores are depleted and short-circuited by applying Ca²⁺ ionophores (ionomycin and A23187), serotonin still induces the normal Ca²⁺ response in the neuronal hybrid cells but not in the glioma cells. Thus, in the neuronal cell line cytosolic Ca²⁺ activity is enhanced through stimulation of Ca²⁺ entry into the cells from the extracellular environment via 5-HT₃ receptors. The depolarization response caused by serotonin in these cells is due to activation of a cation conductance, and consequent entry of extracellular Ca²⁺. In the neuronal cell line, serotonin induces a rise of the cyclic GMP level, which depends on the rise of cytosolic Ca²⁺ activity. This conclusion is derived from the following findings: The serotonin-stimulated rise of cyclic GMP level is inhibited by i) reduced extracellular Ca²⁺ concentration (half-maximal stimulation at 0.3 mM Ca²⁺); ii) addition of inorganic (La³⁺, Mn²⁺, Co²⁺, Ni²⁺) or organic blockers (diltiazem, methoxyverapamil) of Ca²⁺ permeable channels; and iii) intracellular application of the Ca²⁺ chelator BAPTA. The suppression of the cyclic GMP effect of serotonin by the arginine analogues (N^G-monomethyl-L-arginine, N^G-nitro-L-arginine and canavanine) and by incubation in media containing oxyhemoglobin indicates that after stimulation with serotonin nitric oxide released from arginine acts as an intercellular stimulant of soluble guanylate cyclase. The rise of cytosolic Ca²⁺ activity appears to be a prerequisite for the formation of nitric oxide as an activator of guanylate cyclase. In the glial cell line, however, ketanserin-sensitive 5-HT₂ receptors mainly cause liberation of Ca²⁺ from internal stores. In the glioma cells, Ca²⁺ released from internal stores opens Ca²⁺ -dependent K⁺ channels which results in the hyperpolarizing response.