Dynorphin-A-(1-13) antagonizes morphine analgesia in the brain and potentiates morphine analgesia in the spinal cord.

Intrathecal injection of subanalgesic doses of morphine (7.5 nmol) and dynorphin-A-(1-13) (1.25 nmol) in combination resulted in a marked analgesic effect as assessed by tail flick latency in the rat. The analgesic effect of the composite dynorphin/morphine was dose-dependent in serial dilutions so that a composition of 1/8 of the analgesic dose of dynorphin and 1/3 that of morphine produced an analgesic effect equipotent to full dose of either drug applied separately. The analgesic effect induced by dynorphin/morphine mixture was not accompanied by motor dysfunction and was easily reversed by a small dose (0.5 mg/kg) of naloxone. Contrary to the augmentatory effect of dynorphin on morphine analgesia in the spinal cord, intracerevroventricular (ICV) injection of 20 nmol of dynorphin-A-(1-13) exhibited a marked antagonistic effect on the analgesia produced by morphine (120 nmol, ICV). The theoretical considerations and practical implications of the differential interactions between dynorphin-A-(1-13) and morphine in the brain versus spinal cord are discussed.

Mobilization of calcium from intracellular store as a possible mechanism underlying the anti-opioid effect of angiotensin II.

Angiotensin II (AII), injected intracerebroventricularly, has been shown to antagonize opioid analgesia. The mechanism for this was obscure. In the neuroblastoma X glioma NG 108-15 hybrid cell line, the K(+)-induced increase in [Ca2+]i can be suppressed by the delta opioid agonist [D-Pen2, D-Pen5]enkephalin (DPDPE) at 0.01-1 microM, an effect completely reversed by the opioid antagonist naloxone. Angiotensin II (AII) at concentrations of 0.1 and 1 microM mobilized free Ca2+ from an intracellular pool, and this effect was antagonized by the AII receptor antagonist saralasin. All (1 microM) had no significant effect on the increase in [Ca2+]i induced by K+, but it blocked the suppressive effect of DPDPE on the K(+)-induced [Ca2+]i increase. The results indicate that mobilization of intracellular calcium may underlie the anti-opioid effect of AII.

Cholecystokinin-8 suppressed 3H-etorphine binding to rat brain opiate receptors.

Radio receptor assay (RRA) was adopted to analyse the influence of CCK-8 on 3H-etorphine binding to opiate receptors in rat brain synaptosomal membranes (P2). In the competition experiment CCK-8 (1pM to 1 microM) suppressed the binding of 3H-etorphine. This effect was completely reversed by proglumide at 1 microM. Rosenthal analysis for saturation revealed two populations of 3H-etorphine binding sites. CCK-8 (1pM to 1 microM) inhibited 3H-etorphine binding to the high affinity sites by an increase in Kd (up to +235%) and decrease in Bmax (up to -80%) without significant changes in the Kd and Bmax of the low affinity sites. This effect of CCK-8 (10nM) was also completely reversed by proglumide at 1 microM. Unsulfated CCK-8 (100pM to 1 microM) produced only a slight increase in Kd of the high affinity sites (+64%) without affecting Bmax. The results suggest that CCK-8 might be capable of suppressing the high affinity opioid binding sites via the activation of CCK receptor.

Suppression by [D-Pen2, D-Pen5]enkephalin on cyclic AMP dependent protein kinase-induced, but not protein kinase C-induced increment of intracellular free calcium in NG108-15 cells.

In neuroblastoma X glioma NG108-15 cell lines, KCl 50 mM produced a significant increase in [Ca2+]i which was blocked completely by voltage-dependent Ca2+ channel antagonist verapamil. High K(+)-induced increase in [Ca2+]i can be suppressed by selective delta opioid agonist [D-Pen2, D-Pen5]enkephalin (DPDPE) (an effect completely reversed by opioid antagonist naloxone), but not by the mu agonist ohmefentanyl (OMF) or kappa agonist 66A-078. Aside from high K+ stimulation, a number of chemicals can produce an increase in [Ca2+]i, i.e., selective adenylate cyclase activator forskolin, the membrane permeable cyclic AMP (cAMP) analogue dibutyryl-cAMP (Bt2cAMP) and the activator of protein kinase C (PKC) 12-O-tetradecanoylphorbol 13-acetate (TPA). All these effects can be readily blocked by verapamil. DPDPE blocks the increase in [Ca2+]i induced by forskolin and Bt2cAMP, but not that by TPA. The results suggest that cAMP dependent protein kinase-, but not PKC-induced Ca2+ influx mechanism seems to be involved in the delta receptor mediated opioid effect.

Cyclic AMP mediates mu and delta, but not kappa opioid analgesia in the spinal cord of the rat.

Intrathecal (i.t.) injection of the membrane-permeable cyclic AMP (cAMP) analogue dibutyryl-cyclic AMP (Bt2cAMP) in 3 successive doses of 12.5, 12.5 and 25 ug in rats was very effective in reversing the antinociceptive effects produced by mu agonist morphine or delta agonist DPDPE, but not that by kappa agonist dynorphin A-(1-13). cAMP content of the spinal cord was significantly decreased by morphine, but not by dynorphin A-(1-13). The results imply that a decrease in spinal cAMP content may be important for the antinociceptive effect elicited by mu and delta, but not kappa opioid receptor agonists.

Central neurotransmitters and acupuncture analgesia.

The role played by central neurotransmitters in acupuncture analgesia was evaluated by correlating neurochemical changes in central nervous system with the acupuncture effect, as well as modification of the acupuncture effects by pharmacological manipulations of central neurotransmitters. The results of experimental studies which were performed mainly on rats and rabbits indicated that central serotonin and endogenous opiate-like substances (OLS) seem to be the most important substrates for mediation of acupuncture analgesia while central catecholamines, especially norepinephrine through alpha receptors, may exert an antagonistic effect. It was also found that prolonged and repeated acupuncture resulted in a gradual decrease of the acupuncture effects. The development of some endogenous anti-opiate substrates (AOS) in central nervous system was tentatively implicated.

Effect of calcium ion on analgesia of opioid peptides.

The analgesic effect produced by subcutaneous injection (SC) of morphine was antagonized by intracerebroventricular (ICV) but not intrathecal (ITH) injection of CaCl2. While ITH CaCl2 was devoid of any effect on the analgesia induced by ITH morphine, it did antagonize the analgesic effect produced by ITH injection of dynorphin A or (D-Pen2,D-Pen5)-enkephalin (DPDPE). In accordance with this, the uptake of 45Ca by synaptosomes prepared from the dorsal column of rat spinal cord was suppressed by dynorphin A or DPDPE, but not by morphine. The results imply that different kinds of opioid ligands might have different mechanisms of action at discrete areas of the CNS. Blockade of neuronal Ca2+ uptake may serve as an important mechanism for morphine analgesia in the brain as well as DPDPE and dynorphin A analgesia in the spinal cord.