The 5-HT 2A/2C agonist (Ϯ)-1-(2,5-dimethoxy-4-iodophenyl)-2aminopropane HCl (DOI) stimulates hypothalamic neurons to increase the secretion of several hormones. This study addressed two questions: 1) are the neuroendocrine effects of DOI mediated via activation of 5-HT 2A receptors; and 2) which neurons are activated by 5-HT 2A receptors. The 5-HT 2A antagonist (ϩ)-␣-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidinemethanol (MDL 100,907; 0.001, 0.01, or 0.1 mg/kg, s.c.) was administered before rats were challenged with DOI (2.5 mg/kg, i.p.). MDL 100,907 produced a dose-dependent inhibition (ED 50 Х 0.001 mg/kg) of the effect of DOI on plasma levels of ACTH, corticosterone, oxytocin, prolactin, and renin without altering basal hormone levels. Complete blockade of the effect of DOI was achieved for all hormones at MDL 100,907 doses of 0.01-0.1 mg/kg. In a parallel experiment, DOI was injected 2 hr before killing to determine its effects on the expression of Fos, the product of the immediate early gene c-fos. DOI induced an increase in Fos immunoreactivity in corticotropin-releasing factor (CRF) and in oxytocin-expressing neurons but not in vasopressin-containing neurons in the hypothalamic paraventricular nucleus or CRF cells in the amygdala. Pretreatment with MDL 100,907 (0.1 mg/kg, s.c.) blocked the DOI-induced increase in Fos expression in all regions including the hypothalamus, amygdala (central and corticomedial), bed nucleus of the stria terminalis, and prefrontal cortical regions. The combined neuroanatomical and pharmacological observations suggest that the neuroendocrine responses to DOI are mediated by activation of neurons in the hypothalamic paraventricular nucleus and associated circuitry. Furthermore, selective activation of 5-HT 2A receptors mediates the hormonal and Fos-inducing effects of DOI.

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Using in situ hybridization and immunoblot analysis, the present studies identified G z mRNA and G z -protein in the hypothalamic paraventricular nucleus. The role of G z -proteins in hypothalamic 5-HT 1A receptor signaling was examined in vivo. Activation of 5-HT 1A receptors increases the secretion of oxytocin and ACTH, but not prolactin. Intracerebroventricular infusion (3-4 d) of G z antisense oligodeoxynucleotides, with different sequences and different phosphorothioate modification patterns, reduced the levels of G z -protein in the hypothalamic paraventricular nucleus, whereas missense oligodeoxynucleotides had no effect. Neither antisense nor missense oligodeoxynucleotide treatment altered basal plasma levels of ACTH, oxytocin, or prolactin, when compared with untreated controls. An antisense-induced decrease in hypothalamic G z -protein levels was paralleled by a significant decrease in the oxytocin and ACTH responses to the 5-HT 1A agonist 8-hydroxydipropylamino-tetralin (8-OH-DPAT). In contrast, the prolactin response to 8-OH-DPAT (which cannot be blocked by 5-HT 1A antagonists) was not inhibited by G z antisense oligodeoxynucleotides. G z -proteins are the only members of the G i / G o -protein family that are not inactivated by pertussis toxin. In a control experiment, pertussis toxin treatment (1 g/5 l, i.c.v.; 48 hr before the 8-OH-DPAT challenge) did not inhibit the ACTH response, potentiated the oxytocin response, and eliminated the prolactin response to 8-OH-DPAT. Thus, pertussis toxinsensitive G i /G o -proteins do not mediate the 5-HT 1A receptormediated increase in ACTH and oxytocin secretion. Combined, these studies provide the first in vivo evidence for a key role of G z -proteins in coupling hypothalamic 5-HT 1A receptors to effector mechanisms.