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

Abstract

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 (NG-monomethyl-L-arginine, NG-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.