The role of 5-HT2A-mediated stimulation of phosphoinositide hydrolysis in the discriminative effects of hallucinogens was investigated in PC12 cells stably expressing the rat 5-HT2A receptor (PC12-5-HT2A 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 beta-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-HT2A-mediated stimulation of phosphoinositide hydrolysis does not appear to be the sole critical signaling mechanism involved in the discriminative effects of hallucinogens.