A-381393

In-vitro studies have shown that WAY 100635 is not only a potent 5-HT1A antagonist, but also has high affinity and efficacy at the dopamine D(4) receptor. Nevertheless, the behavioral effects of this compound have not been investigated. This study sought to characterize the discriminative stimulus effects produced by WAY 100635. Male Sprague-Dawley rats were trained in a two-lever, fixed ratio 50, food-reinforced task with WAY 100635 (10 micromol/kg) as a discriminative stimulus. Substitution tests with different doses of WAY 100635 and (-)-pindolol, and combination tests with two 5-HT1A agonists, 8-OH-DPAT and LY 293284; and two dopamine D(4) antagonists, sonepiprazole and A-381393, were performed. Rats trained with a low dose (0.74 micromol/kg) of WAY 100635 could not learn the discrimination task after more than 3 months of sessions. Rats trained to discriminate 10 micromol/kg of WAY 100635 from saline achieved the criterion of accuracy after approximately 35 training sessions. WAY 100635 (2.5-10 micromol/kg) produced a dose-dependent increase in WAY 100635-appropriate responding, with a mean effective dose of 3.44 micromol/kg, whereas saline or pindolol (5-25 micromol/kg) administration resulted in 0% drug lever responding. Pretreatment with the 5-HT(1A) agonists 8-OH-DPAT or LY 293284 did not modify the WAY 100635 curve, but pretreatment with the selective dopamine D(4) antagonists sonepiprazole or A-381393 completely blocked the cue. These results indicate that the discriminative stimulus effect produced by WAY 100635 is mediated by activation of dopamine D(4) receptors.

The dopamine D4 receptor has been investigated for its potential role in several CNS disorders, notably schizophrenia and more recently, erectile dysfunction. Whereas studies have investigated dopamine D4 receptor-mediated signaling in vitro, there have been few, if any, attempts to identify dopamine D4 receptor signal transduction pathways in vivo. In the present studies, the selective dopamine D4 agonist PD168077 induces c-Fos expression and extracellular signal regulated kinase (ERK) phosphorylation in the hypothalamic paraventricular nucleus (PVN), a site known to regulate proerectile activity. The selective dopamine D4 receptor antagonist A-381393 blocked both c-Fos expression and ERK1/2 phosphorylation produced by PD168077. In addition, PD168077-induced ERK1/2 phosphorylation was prevented by SL327, an inhibitor of ERK1/2 phosphorylation. Interestingly, treatment with A-381393 alone significantly reduced the amount of Fos immunoreactivity as compared to basal expression observed in vehicle-treated controls. Dopamine D4 receptor and c-Fos coexpression in the PVN was observed using double immunohistochemical labeling, suggesting that PD168077-induced signaling may result from direct dopamine D4 receptor activation. Our results demonstrate functional dopamine D4 receptor expression and natural coupling in the PVN linked to signal transduction pathways that include immediate early gene and MAP kinase activation. Further, the ability of the selective dopamine D4 antagonist A-381393 alone to reduce c-Fos expression below control levels may imply the presence of a tonic dopamine D4 receptor activation under basal conditions in vivo. These findings provide additional evidence that the PVN may be a site of dopamine D4 receptor-mediated proerectile activity.