5-HT_2A receptor-stimulated phosphoinositide hydrolysis in the stimulus effects of hallucinogens

Abstract

The role of 5-HT_2A-mediated stimulation of phosphoinositide hydrolysis in the discriminative effects of hallucinogens was investigated in PC12 cells stably expressing the rat 5-HT_2A receptor (PC12-5-HT_2A cells). The hallucinogenic compounds, d-lysergic acid diethylamide (LSD), (−)2,5-dimethoxy-4-methylamphetamine (DOM), psilocybin, N,N-dimethyltryptamine (DMT), 5-methoxy-N,N-dimethyltryptamine (MDMT) and N,N-diethyltryptamine (DET), all caused a concentration-dependent increase in the generation of [³H]inositol phosphates. The nonhallucinogenic compounds, 6-fluoro-N,N-diethyltryptamine (6-F-DET), lisuride and quipazine, also displayed significant efficacy in stimulating phosphoinositide hydrolysis, while 2-bromo-lysergic acid diethylamide (BOL), which is not a hallucinogen, did not alter inositol phosphate generation. The β-carbolines, harmaline and harmane, also did not alter phosphoinositide hydrolysis. Comparison of these results with previous drug discrimination studies indicated the apparent lack of correlation between the degree of substitution in LSD- and DOM-trained animals and efficacy in stimulating phosphoinositide hydrolysis. The present study indicates that 5-HT_2A-mediated stimulation of phosphoinositide hydrolysis does not appear to be the sole critical signaling mechanism involved in the discriminative effects of hallucinogens.

Introduction

The serotonergic receptor system appears to play a prominent role in mediating the discriminative effects of both indoleamine [e.g., d-lysergic acid diethylamide (LSD)] and phenethylamine [e.g., (−)2,5-dimethoxy-4-methylamphetamine (DOM)] hallucinogens. The initial finding that serotonergic antagonists block the stimulus effects of the phenethylamine hallucinogen mescaline Winter, 1975, Browne and Ho, 1975 was later expanded to include other hallucinogens including LSD, DOM and N,N-dimethyltryptamine (DMT) Kuhn et al., 1997, Winter, 1978, Glennon et al., 1983. Of the seven different families of serotonin receptors, activation of the 5-HT₂ receptor family appears to be the primary site of action of the indoleamine/phenethylamine hallucinogens. Affinity at the 5-HT₂ receptor for a series of phenethylamines strongly correlates with both the potency of these compounds to substitute for DOM as a discriminative cue (Glennon et al., 1984), as well as their potency as hallucinogens Titeler et al., 1988, Sadzot et al., 1989. Based upon antagonist correlation analysis, Fiorella et al. (1995a) concluded that the 5-HT_2A receptor rather than the 5-HT_2C receptor mediates the stimulus effects of LSD. Similarly, the discriminative stimulus effects of the phenethylamine DOI are blocked by MDL 100,907, a selective 5-HT_2A antagonists, but not by the selective 5-HT_2C antagonist SB 200,646 (Schreiber et al., 1994). In addition, behavioral tolerance to the stimulus effects of DOI is associated with a down-regulation of the 5-HT_2A and not the 5-HT_2C receptor (Smith et al., 1999).

Although the data indicate that the discriminative stimulus effects of indoleamine and phenethylamine hallucinogens involve activation of the 5-HT_2A receptor, the resulting biochemical changes responsible for the interceptive state induced by these hallucinogens are unknown. The 5-HT_2A receptor is coupled to stimulation of phospholipase C through an activation of the G_q/11 GTP-binding protein. This stimulation results in the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP₂) with the subsequent generation of inositol 1,4,5-trisphosphate (IP₃), which regulates the release of calcium from internal stores, and sn(1,2) diacylglycerol (DAG), which is involved in the activation of protein kinase C. The objective of the present study was to determine whether 5-HT_2A receptor-mediated stimulation of phosphoinositide hydrolysis is a necessary component of the discriminative state produced by hallucinogens. To address this question, generation of inositol phosphates was measured in the presence of various indoleamine, phenethylamine and β-carboline hallucinogens, as well as some nonhallucinogenic congeners. For these studies, PC12 cells expressing the rat 5-HT_2A receptor cDNA were used to preclude the confounding stimulation of phospholipase C by other members of the 5-HT₂ receptor family.