The spontaneously hypertensive (SHR) and Lewis (LEW) strains differ in numerous behavioral tests, including the elevated plus-maze. In keeping with the crucial role of central serotonin (5-HT) in anxiety, we checked for strain differences regarding several determinants of 5-HT activity. In addition to confirming that LEW rats displayed anxious behaviors in the plus-maze compared with SHR, we found that in vitro, central tryptophan hydroxylase activity was higher in LEW rats than in SHR. However, ex vivo studies in midbrains and hippocampi revealed that neither 5-HT synthesis nor 5-HT and 5-hydroxyindoleacetic acid levels differed between strains. [3H]8-Hydroxy-2-(di-n-pro-pylamino)tetralin binding at midbrain 5-HT1A autoreceptors and hippocampal 5-HT1A postsynaptic receptors, [3H]ketanserin binding at cortical and striatal 5-HT2A receptors and [3H]citalopram binding at midbrain and hippocampal 5-HT transporters did not vary between strains. The inhibition of 5-HT synthesis by 5-HT1A autoreceptor stimulation was similar in both strains. Forepaw treading and flat body posture after 5-HT1A postsynaptic receptor stimulation were higher and lower, respectively, in SHR than in LEW rats. Last, 1-(4-iodo-2,5-dimethoxy-phenyl)-2-aminopropane- and quipazine-elicited head shakes, a 5-HT2A receptor-mediated response, were increased in the SHR strain compared with the LEW strain; on the other hand, 1-(3-chlorophenyl)piperazine triggered similar 5-HT2B/2C receptor-mediated decreases in motor activity in the two strains. This study shows that although the low-anxiety (SHR) and high-anxiety (LEW) strains vary in some aspects of 5-HT function, key components such as the 5-HT1A autoreceptors are not different.