Modulation of adenylate cyclase activity of mouse spinal cord-ganglion explants by opioids, serotonin and pertussis toxin

Abstract

Organotypic cultures of fetal mouse spinal cord-ganglion explants (2–4 weeks in vitro) contain forskolin-stimulated adenylate cyclase (AC) activity that is inhibited by levorphanol and other opioid agonists in a dose-dependent manner. Inhibition by levorphanol no longer occurs if sodium is omitted from the incubation and the levorphanol inhibition is blocked by the opioid antagonist, naloxone. These findings together with the ineffectiveness of dextrophan indicate that the opioid inhibition of forskolin-stimulated AC is receptor mediated. Both the δ- and κ-receptor subtypes appear to be involved since the selective δ-opioid agonist, [ d-Pen 2. d-Pen 5]enkephalin, and the selective κ-opioid agonist, t-3,4-dichloro- N-methyl- N[2-(1-pyrrolidinyl)cyclohexyl]-benzene acetamide (U-50,488H) are both effective at nanomolar concentrations. In contrast, the selective μ-opioid agonists, Tyr- d-Ala-Gly -N- MePhe-Gly-ol , has no significant effect even at micromolar concentrations. Both cord and ganglion components of the explants contain opioid-sensitive AC. Forskolin-stimulated AC of the explants is also inhibited by serotonin and carbachol. The serotonin effect appears to be mediated by 5-HT 1A receptors, based on relative agoonist and antagonist selectivity. Chronic exposure of cultures to morphine results in enhanced basal and forskolin-stimulated AC as well as attenuation of opioid-inhibition of AC assayed in the presence of forskolin: treatment of explants with pertussis toxin causes similar changes in the AC system. The inhibitory effect of serotonin is also attenuated by the pertussis toxins treatment. Basal AC activity of the explants (assayed without forskolin present) is stimulated to a small but significant extent opioids and by serotonin. The opioid stimulatory effect is markedly enhanced following either morphine or pertussis toxin treatment of the explants. The attenuation of opioid- and serotonin-inhibition of AC produced by chronic exposure to pertussis toxin and the attenuation of opioid inhibition produced by exposure to morphine are consonant with the attenuation of opioid and monoaminergic depression of sensory evoked dorsal horn network responses after similar chronic treatments. It is proposed that the inhibitory effects of opioids and seroronin on these neurons are mediated by receptors that are negatively coupled via a pertussis toxin sensitive G 1 protein to AC. Furthermore, alterations of AC with chronic morphine treatment may be involved in the development of physiologic tolerance to opioids.