Circadian rhythms have been demonstrated in mammals for a variety of biological phenomena, including running activity,☂ rectal temperature,? plasma eosinophil and corticosterone levels,?: * liver mitoses, and nucleic acid content.☁ Certain of these rhythmsare considered endogenous since they persist in the absence of environmental lighting.!:* Controlling centers within the organism for these rhythms have been sought without success. Thus adrenal-dependent rhythms in epidermal mitoses and rectal temperature persist after hypophysectomy, although their am-

Rat adrenal gland levels of dopamine &hydroxylase are subject to dual control. Activation of the splanchnic nerves to the adrenal medulla by reserpine induces the synthesis of dopamine /3-hydroxylase without altering the rate of enzyme degradation. In contrast, hypophysectomy causes a decline in steady state dopamine fi-hydroxylase levels by first accelerating the rate of degradation, then by slowing the rate of enzyme synthesis as well. Adrenocorticotropic hormone administration partially reversed the effect of hypophysectomy on dopamine B-hydroxylase degradation. These findings suggest that the trans-synaptic factors controlling dopamine P-hydroxylase induction act by a different mechanism (enzyme synthesis) than the hormonal controls regulating steady state levels (enzyme degradation). Thus, active inhibition of enzyme degradation may be an important control in maintenance of steady state enzyme levels.

A bstract : Cannabidiol and other cannabinoids were examined as neuroprotectants in rat cortical neuron cultures exposed to toxic levels of the neurotransmitter, glutamate. The psychotropic cannabinoid receptor agonist Δ 9 ‐tetrahydrocannabinol (THC) and cannabidiol, (a non‐psychoactive constituent of marijuana), both reduced NMDA, AMPA and kainate receptor mediated neurotoxicities. Neuroprotection was not affected by cannabinoid receptor antagonist, indicating a (cannabinoid) receptor‐independent mechanism of action. Glutamate toxicity can be reduced by antioxidants. Using cyclic voltametry and a fenton reaction based system, it was demonstrated that Cannabidiol, THC and other cannabinoids are potent antioxidants. As evidence that cannabinoids can act as an antioxidants in neuronal cultures, cannabidiol was demonstrated to reduce hydroperoxide toxicity in neurons. In a head to head trial of the abilities of various antioxidants to prevent glutamate toxicity, cannabidiol was superior to both a‐tocopherol and ascorbate in protective capacity. Recent preliminary studies in a rat model of focal cerebral ischemia suggest that cannabidiol may be at least as effective in vivo as seen in these in vitro studies.

The neuroprotective actions of cannabidiol and other cannabinoids were examined in rat cortical neuron cultures exposed to toxic levels of the excitatory neurotransmitter glutamate. Glutamate toxicity was reduced by both cannabidiol, a nonpsychoactive constituent of marijuana, and the psychotropic cannabinoid (-)Delta9-tetrahydrocannabinol (THC). Cannabinoids protected equally well against neurotoxicity mediated by N-methyl-D-aspartate receptors, 2-amino-3-(4-butyl-3-hydroxyisoxazol-5-yl)propionic acid receptors, or kainate receptors. N-methyl-D-aspartate receptor-induced toxicity has been shown to be calcium dependent; this study demonstrates that 2-amino-3-(4-butyl-3-hydroxyisoxazol-5-yl)propionic acid/kainate receptor-type neurotoxicity is also calcium-dependent, partly mediated by voltage sensitive calcium channels. The neuroprotection observed with cannabidiol and THC was unaffected by cannabinoid receptor antagonist, indicating it to be cannabinoid receptor independent. Previous studies have shown that glutamate toxicity may be prevented by antioxidants. Cannabidiol, THC and several synthetic cannabinoids all were demonstrated to be antioxidants by cyclic voltametry. Cannabidiol and THC also were shown to prevent hydroperoxide-induced oxidative damage as well as or better than other antioxidants in a chemical (Fenton reaction) system and neuronal cultures. Cannabidiol was more protective against glutamate neurotoxicity than either ascorbate or alpha-tocopherol, indicating it to be a potent antioxidant. These data also suggest that the naturally occurring, nonpsychotropic cannabinoid, cannabidiol, may be a potentially useful therapeutic agent for the treatment of oxidative neurological disorders such as cerebral ischemia.

In this unpublished manuscript, Axelrod wrote on signal transduction in neurotransmitters, some of which was based on his work as a guest researcher at the NIMH in the Laboratory of Cell Biology in the late 1980s and early 1990s.

Arginine vasopressin mediates its effects through vasopressin receptor activation and second messenger production. Recent cloning of the V1a receptor provided the opportunity to investigate the possible signal transduction pathways associated with this single vasopressin receptor subtype. When stably expressed in CHO cells, vasopressin stimulated several signal transduction pathways simultaneously including calcium influx, phospholipase A2, phospholipase C, and phospholipase D. Vasopressin-stimulated release of arachidonic acid, IP3 formation, and phosphatidylethanol formation (in the presence of 1% ethanol) were used as indexes of phospholipase A2, phospholipase C, and phospholipase D activation, respectively. V1a receptor-activation stimulated a peak followed by a sustained plateau phase of intracellular calcium. The plateau phase was dependent on extracellular calcium, insensitive to blockers of voltage sensitive calcium channels, blocked by heavy metals, and quenched when MnCl2 was present in the extracellular media. Removal of extracellular calcium blunted the release of IP3, and blocked the release of arachidonic acid and phosphatidylethanol indicating that these responses were at least in part regulated by receptor-operated calcium influx. Vasopressin-stimulated release of arachidonic acid and phosphatidylethanol were augmented with the phorbol ester PMA, and this augmentation was blocked by inhibitors of protein kinase C and absent with long-term PMA treatment. Vasopressin-stimulated IP3 release was inhibited with PMA and the inhibition reversed with protein kinase C inhibitors.

An enzyme system in rabbit liver microsomes which catalyzes the demethylation of ephedrine to yield norephedrine and formaldehyde is described. The enzyme system requires reduced triphosphopyridine nucleotide and oxygen. Species differences in the metabolism of ephedrine may be explained in part by differences in the activity of ephedrine-demethylating enzyme and in part by the presence of a heat labile inhibitory factor in microsomes. A heat stable factor which can stimulate the enzymatic demethylation of ephedrine is also present in microsomes.

ATP is copackaged and coreleased with adrenergic, serotonergic, and cholinergic neurotransmitters, suggesting a possible interaction between the signaling pathways for ATP and these coreleased neurotransmitters. Muscarinic m2 and m4, alpha-2-adrenergic, and D2-dopaminergic neurotransmitter receptors, which have in common their ability to inhibit adenylate cyclase through the inhibitory guanine nucleotide binding protein G(i), were transfected and expressed in Chinese hamster ovary (CHO) cells that contain endogenous ATP receptors coupled to the release of arachidonic acid. Normal functional coupling of m2, m4, alpha-2, and D2 receptors was demonstrated by their ability to inhibit forskolin-stimulated cAMP accumulation with dose-response activities consistent with previous reports for these G(i)-coupled receptors. Stimulation of m2, m4, alpha-2, and D2 receptors resulted in an augmentation of ATP-stimulated arachidonic acid release. With the exception of the m4 receptor, none of the receptors tested was able to stimulate arachidonic acid release in the absence of ATP. Potentiation of ATP-stimulated arachidonic acid release was independent of changes in cAMP. The augmentation of ATP-stimulated arachidonic acid release and the inhibition of cAMP accumulation were both blocked by pertussis toxin, an inhibitor of G(i), but with different dose-response characteristics. Inhibition of protein kinase C with staurosporine or long-term pretreatment of the cells with the phorbol ester phorbol 12-myristate 13-acetate blocked the augmentation response. This demonstrates that G(i)-coupled inhibitory receptors can amplify ATP-receptor-stimulated arachidonic acid release through a pertussis-toxin-sensitive G protein, independent of their ability to inhibit adenylate cyclase activity.

1. This study demonstrates that human recombinant interleukin-1 (IL-1) stimulates beta-endorphin release and potentiates the secretion of beta-endorphin in both a mouse anterior pituitary cell line AtT-20 and rat pituitary cell cultures. 2. In pituitary cell cultures, prolonged treatment with phorbol ester had no effect on IL-1-induced beta-endorphin release, but abolished the potentiating effects of IL-1 on vasopressin-induced beta-endorphin secretion. 3. The enhancement of CRF-stimulated beta-endorphin release by IL-1 was also reduced in normal pituitary cell cultures following depletion of protein kinase C. 4. The late IL-1-induced secretion of beta-endorphin does not require the continuous presence of the cytokine. 5. Incubation of monolayers with 125I-IL-1 alpha (10(-9) M) at 8 degrees C and then at 37 degrees C for various times revealed that IL-1 alpha was internalized. There was a progressive increase in the ratio of cytoplasmic to cell-surface-associated 125I-IL-1 alpha. 6. These results indicate that the IL-1-induced beta-endorphin release and its potentiation of beta-endorphin secretion involves internalization of this cytokine, perhaps via cell surface IL-1 receptors.

A rat D2L dopamine receptor, a splice variant of the D2 receptor, has recently been cloned. When transfected into and stably expressed in Chinese hamster ovary cells, these receptors mediate the inhibition of both basal and forskolin-stimulated cAMP production, as previously described. We examined what role this receptor might play in the production of the second messenger arachidonic acid. The calcium ionophore A23187 stimulated the release of arachidonic acid, and this release of arachidonic acid was potentiated by dopamine in a concentration-dependent manner. Dopamine alone, however, had no effect on arachidonic acid release. Quinpirole, a D2-selective agonist, augmented A23187-stimulated arachidonic acid release, and sulpiride, a D2-selective antagonist, blocked this augmentation. cAMP analogs and agents that activate adenylyl cyclase were utilized in an attempt to overcome this dopamine effect. Forskolin, prostaglandin E2, dibutyryl-cAMP, 8-(4-chlorophenylthio)-cAMP, and pertussis toxin all had no appreciable effect on either A23187-stimulated arachidonic acid release or the dopamine enhancement. Inhibition of protein kinase C using long term phorbol ester desensitization and pharmacological inhibitors diminished the dopamine potentiation of arachidonic acid release. These results suggest that the D2 receptor may be increasing the release of arachidonic acid by a mechanism involving protein kinase C but independent of the D2 receptor's inhibition of adenylyl cyclase.

Interleukin 1 (IL-1) has been shown to potentiate the release of beta-endorphin induced by secretagogues, including corticotropin releasing factor (CRF) and phorbol ester (TPA), in the mouse AtT-20 pituitary tumor cell line (Fagarasan et al., PNAS, 1989, 86, 2070-2073). In cultured rat anterior pituitary cells, pretreatment with IL-1 caused only a small increase in beta-endorphin release but significantly potentiated CRF-and vasopressin-stimulated beta-endorphin secretion. Vasopressin stimulates the secretion of beta-endorphin in normal pituitary cells but not in AtT-20 cells. However, treatment of AtT-20 cells with IL-1 induced the expression of vasopressin-mediated beta-endorphin release; this effect of IL-1 was reduced after depletion of protein kinase C by prolonged treatment with TPA. The enhancement of CRF-stimulated beta-endorphin release by IL-1 was also reduced in AtT-20 cells after depletion of protein kinase C, and after treatment with staurosporine. These findings indicate that treatment with IL-1 amplifies receptor-mediated responses to the major physiological secretagogues in normal corticotrophs, and initiates a secretory response to vasopressin in AtT-20 cells.

Conference Article| August 01 1990 Receptor-mediated activation of phospholipase A2 and arachidonic acid release in signal transduction JULIUS AXELROD JULIUS AXELROD 1Laboratory of Cell Biology, National Institute of Mental Health, Bethesda, MD 20892, U.S.A. Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (1990) 18 (4): 503–507. https://doi.org/10.1042/bst0180503 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation JULIUS AXELROD; Receptor-mediated activation of phospholipase A2 and arachidonic acid release in signal transduction. Biochem Soc Trans 1 August 1990; 18 (4): 503–507. doi: https://doi.org/10.1042/bst0180503 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Society Transactions Search Advanced Search This content is only available as a PDF. © 1990 Biochemical Society1990 Article PDF first page preview Close Modal You do not currently have access to this content.

Serotonin (5-HT) stimulated the release of arachidonic acid in hippocampal neurons cocultured with glial cells but not in glial cultures alone. Similar results were observed for the 5-HT-stimulated release of inositol phosphates. These results suggest a neural but not glial origin of both responses. Pharmacological studies suggested that release of arachidonic acid and inositol phosphates was mediated by a type 2 5-HT (5-HT2) receptor. 5-HT-stimulated release of arachidonic acid was also detected in cortical neurons, which contain high levels of 5-HT2 receptors, but not striatum, spinal cord, or cerebellar granule cells, which have very low levels or are devoid of 5-HT2 receptors. The phorbol ester phorbol 12-myristate 13-acetate augmented the 5-HT-stimulated release of arachidonic acid but inhibited the 5-HT-stimulated release of inositol phosphates. 5-HT-stimulated release of arachidonic acid, but not inositol phosphates, was dependent on extracellular calcium. 5-HT stimulated the release of [3H]lysophosphatidylcholine from [3H]choline-labeled cells with no increase in the release of [3H]choline or phospho[3H]choline. These data suggest that 5-HT stimulated the release of arachidonic acid in hippocampal neurons through the activation of phospholipase A2, independent of the activation of phospholipase C.