E. L. Way

Opiate binding to cerebroside sulfate: a model system for opiate-receptor interaction.

Abstract Cerebroside sulfate was shown to bind etorphine and levorphanol with high affinity. The relative potency of narcotic analgesics in preventing the binding of levorphanol to cerebroside sulfate correlated well with their reported analgetic activity. The data indicate similarities between cerebroside sulfate and a purified opiate receptor from mouse brain which has been reported to be a proteolipid. Some preliminary animal data also imply the involvement of CS in opiate action We, therefore, propose that CS may serve as a useful “receptor” model for the study of opiate-receptor interaction in vitro .

Effects of morphine tolerance and dependence on Mg++ dependent ATPase activity of synaptic vesicles.

Abstract The effect of morphine on ATPase of synaptic plasma membranes (SPM) and synaptic vesicles isolated from the mouse brain was studied. The activity of synaptic vesicle Mg ++ -dependent ATPase from mice rendered morphine tolerant and dependent by pellet implantation was 40% higher than that from placebo implanted mice. However, the activities of Mg ++ -dependent ATPase and Na + , K + activated ATPase of SPM of tolerant and nontolerant mice were not significantly different. The activity of synaptic vesicular Mg ++ -dependet ATPase was dependent on the concentration of Mg ++ but not of Ca ++ ; maximum activity was obtained with 2 mM MgCl 2 . On the other hand, Mg ++ -dependent ATPase activity of SPM was dependent on both Mg ++ and Ca ++ , activity being maximum using 2 mM MgCl 2 and 10 −5 M CaCl 2 . It is suggested that this stimulation of ATPase activity may alter synaptic transmission and may thus be involved in some aspects of morphine tolerance and dependence.

Application of an irreversible opiate antagonist (β-FNA, β-funal-trexamine) to demonstrate dynorphin selectivity for K-opioid sites

Abstract Application of 100 nM β-FNA for 60 minutes to isolated longitudinal muscles-myenteric plexus preparations from the guinea pig ileum caused a marked antagonism of the inhibitory action of normorphine and leucine enkephalin without greatly affecting the inhibitory potency of dynorphin or ethylketocyclazocine. The interaction of β-FNA with the normorphine (μ-opiate receptors) appears to be non-equilibrium. Pretreatment with β-FNA caused a significant increase in the apparent naloxone dissociation constant for normorphine and leucine enkephalin but not for dynorphin or ethylketocyclazocine. The results lend further support to the hypothesis that normorphine and the enkephalins activate preferentially μ-opiate receptors on the ileum, whereas dynorphin interacts predominantly at k-opiate sites.

Enhancement of pentobarbital effect by continuous administration of morphine in the mouse

Abstract These studies demonstrated that continuous morphine treatment from implantation of a 75 mg morphine pellet for 3 days potentiated pentobarbital narcosis and enhanced pentobarbital hypothermia. In the morphine implant mice, sleeping time after two different doses of pentobarbital was greater than 2.5 × the sleeping time in placebo pellet implant animals and also greater than sleeping time in animals treated acutely with morphine prior to pentobarbital. Moreover, in the morphine implant mice both the degree and duration of pentobarbital induced hypothermia were enhanced. The above findings were due to slower rate of metabolism of pentobarbital as evidenced by inhibition of hepatic N-demethylation, and higher levels of brain and serum pentobarbital in the morphine implant mice compared to both placebo and acute morphine mice.

Opioid properties of β-lipotropin fragment 60-65, HArgTryGlyGlyPheMetOH

The pharmacologic activity of the hexapeptide fragment corresponding to the amino acid fragment 60–65 in β-lipotropin, (β-LPH-(60–65)) was studied in vitro and in vivo. In binding assays on synaptosomal plasma membrane the peptide was found to be equipotent to met-enkephalin, but behaved differently to cations; in contrast to met-enkephalin both Mn+2 and Na+ enhanced the binding of β-LPH-(60–65) to synaptosomal plasma membrane. On both the quinea pig ileum and mouse vas deferens β-LPH-(60–65) inhibited contractions elicited by electrical stimulation and each effect was reversible by naloxone. On the guinea pig ileum β-LPH-(60–65) was equipotent to met-enkephalin and 0.5 as potent as normorphine but on the vas deferens it was 4.6 times more potent than normorphine. The activities of β-LPH-(60–65) appear to be due to the intact compound rather than to its conversion to met-enkephalin, since the peptide extracted from the ileum assay was found to behave identically as β-LPH-(60–65) with high pressure liquid chromatography. When β-LPH-(60–65) was administered centrally to mice and rats, no overt central actions were observed and an antinociceptive effect could not be demonstrated. Nor did β-LPH-(60–65) antagonize morphine action or precipitate the withdrawal syndrome in morphine dependent animals. It is concluded that the good agreement which generally exists between in vitro and in vivo assay procedures for opiate-like activity of morphine and its surrogates does not necessarily hold for the endogenous peptides with similar actions.

EFFECT OF CALCIUM AND LANTHANUM IN MORPHINE ANALGESIA AND TOLERANCE

Abstract A series of experiments were initiated to study the influence of calcium on morphine action. The analgetic effect of morphine in non-tolerant and tolerant mice was found to be antagonized by intraventricular calcium and enhanced by the cation chelator EGTA. Elevation of brain bioamines by monoamine oxidase inhibition (pargyline) or reducing brain catecholamine by chemical sympathectomy (6-hydroxydopamine) failed to alter the calcium effect. Enhancing calcium influx with the ionophore X537A also decreased morphine analgesia while inhibition of calcium binding to and movement across cell membrane by lanthanum greatly augmented morphine analgesia. Interestingly, lanthanum per se produced analgesia in both mice and rats and the action was inhibited by the narcotic antagonist, naloxone. Moreover, the analgetic sensitive sites in the CNS to lanthanum and morphine were both found to be in the periventricular and periaqueductal gray areas. Cross tolerance between lanthanum and morphine was noted and lanthanum attenuated the withdrawal syndrome in morphine-dependent mice. The development of tolerance to morphine was accompanied by an increased level of calcium in synaptosomes and a decrease in high affinity calcium binding to the synaptosomal plasma membrane fraction but not to the other membrane fractions studied. The effects suggest that chronic morphine treatment decreases the availability of calcium binding sites on the inner surface of the synaptosomal membrane.