dizocilpine-maleate and chelerythrine

The arcuate nucleus (ARC) of the hypothalamus plays a key role in pain processing. Although it is well known that inhibition of NMDA receptor (NMDAR) in ARC attenuates hyperalgesia induced by peripheral inflammation, the underlying mechanism of NMDAR activation in ARC remains unclear. Protein kinase C (PKC) is involved in several signalling cascades activated in physiological and pathological conditions. Therefore, we hypothesised that upregulation of PKC activates NMDARs in the ARC, thus contributing to inflammatory hyperalgesia. Intra-ARC injection of chelerythrine (CC), a specific PKC inhibitor, attenuated complete Freund's adjuvant (CFA) induced thermal and mechanical hyperalgesia in a dose-dependent manner. In vivo extracellular recordings showed that microelectrophoresis of CC or MK-801 (a NMDAR antagonist) significantly reduced the enhancement of spontaneous discharges and pain-evoked discharges of ARC neurons. In addition, CFA injection greatly enhanced the expression of total and phosphorylated PKCγ in the ARC. Interestingly, CFA injection also remarkably elevated the level of phosphorylated NR2B (Tyr1472) without affecting the expression of total NR2B. Importantly, intra-ARC injection of CC reversed the upregulation of phosphorylated NR2B subunits in the ARC. Taken together, peripheral inflammation leads to an activation of NMDARs mediated by PKC activation in the ARC, thus producing thermal and mechanical hyperalgesia.

Topics: Animals; Arcuate Nucleus of Hypothalamus; Behavior, Animal; Benzophenanthridines; Disease Models, Animal; Dizocilpine Maleate; Evoked Potentials; Freund's Adjuvant; Hyperalgesia; Male; Neurons; Phosphorylation; Protein Kinase C; Protein Subunits; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Signal Transduction; Stress, Mechanical; Up-Regulation

Incubation of rat lymphocytes with homocysteine (HC) or homocysteic acid (HCA) was found to increase the stationary levels of free radicals in lymphocytes, the effect of both ligands being mediated by ionotropic receptors activated by N-methyl-D-aspactic acid (NMDA), the expression of which on rat lymphocyte membranes was earlier demonstrated. In agreement with these data, increase of free radicals in the lymphocyte cytoplasm is preceded by an increase in the intracellular calcium levels, activation of protein kinase C, nicotinamide adenine dinucleotide phosphate oxidase and/or nitric oxide synthase. Both HC and HCA increase the production of IFN-γ and TNF-α by lymphocytes and antagonist of NMDA receptors; MK-801 prevents this effect. The data presented show that rat lymphocyte membrane contains functionally active NMDA receptors, which regulate cytokine accumulation.

Topics: Animals; Benzophenanthridines; Calcium; Cell Membrane; Cytokines; Cytoplasm; Dizocilpine Maleate; Flow Cytometry; Fluorescence; Free Radicals; Homocysteine; Interferon-gamma; Lymphocytes; N-Methylaspartate; NADPH Oxidases; Nitric Oxide; Protein Kinase C; Rats; Receptors, N-Methyl-D-Aspartate; Tumor Necrosis Factor-alpha

Previous evidence demonstrates that low dose morphine systemic administration induces acute thermal hyperalgesia in normal mice through microOR stimulation of the inositol signaling pathway. We investigated the site of action of morphine and the mechanism of action of microOR activation by morphine to NMDA receptor as it relates to acute thermal hyperalgesia. Our experiments show that acute thermal hyperalgesia is blocked in periaqueductal gray with the microOR antagonist CTOP, the NMDA antagonist MK801 and the protein kinase C inhibitor chelerythrine. Therefore, a site of action of systemically administered morphine low dose on acute thermal hyperalgesic response appears to be located at the periaqueductal gray. At this supraspinal site, microOR stimulation by systemically morphine low dose administration leads to an increased phosphorylation of specific subunit of NMDA receptor. Our experiments show that the phosphorylation of subunit 1 of NMDA receptor parallels the acute thermal hyperalgesia suggesting a role for this subunit in morphine-induced hyperalgesia. Protein kinase C appears to be the key element that links microOR activation by morphine administration to mice with the recruitment of the NMDA/glutamatergic system involved in the thermal hyperalgesic response.

Topics: Alkaloids; Analysis of Variance; Animals; Behavior, Animal; Benzophenanthridines; Dizocilpine Maleate; Dose-Response Relationship, Drug; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Hyperalgesia; Male; Mice; Morphine; Motor Activity; Pain Measurement; Periaqueductal Gray; Psychomotor Performance; Reaction Time; Receptors, N-Methyl-D-Aspartate; Receptors, Opioid, mu; Somatostatin

Na/K-ATPase prepared from cerebellum granule cells of 10-12-day-old mice is inhibited by glutamate and its agonists, NMDA (ligand for ionotropic receptors) and ACPD (ligand for metabotropic receptors). The inhibition is specific and prevented by subsequent antagonists (MK-801 for ionotropic NMDA-receptors and MCPG for metabotropic receptors). The inhibiting effect of NMDA is significantly reversed by cysteine and that of ACPD by chelerythrine or indolyl maleimide. It is concluded that ionotropic receptors inhibit Na/K-ATPase because of intracellular production of reactive oxygen species, and metabotropic receptors mediate their effect via protein kinase C.

Topics: Alkaloids; Animals; Benzoates; Benzophenanthridines; Cerebellum; Cycloleucine; Cysteine; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Glutamates; Glycine; Male; Mice; Mice, Inbred AKR; N-Methylaspartate; Neurons; Phenanthridines; Receptors, Glutamate; Sodium-Potassium-Exchanging ATPase

In evaluating mechanisms of trimethyltin (TMT)-initiated neuronal damage, the present study focused on involvement of reactive oxygen species, protein kinase C (PKC), and glutamate receptors. Exposure of cerebellar granule cells to TMT (0.01-0.1 microM) produced primarily apoptosis, but higher concentrations were associated with cellular lactate dehydrogenase efflux and necrosis. TMT increased generation of cellular reactive oxygen species, which was inhibited by either L-NAME (inhibitor of nitric oxide synthase, NOS) or catalase, indicating that both NO and H(2)O(2) are formed on TMT exposure. Since chelerythrine (selective PKC inhibitor) also inhibited oxidative species generation, PKC appears to play a significant role in TMT-induced oxidative stress. The metabotropic glutamate receptor antagonist, MCPG, (but not MK-801) prevented oxidative species generation, indicating significant involvement of metabotropic receptors (but not NMDA receptors) in TMT-induced oxidative stress. NOS involvement in the action of TMT was confirmed through measurement of nitrite, which increased concentration dependently. Nitrite accumulation was blocked by L-NAME, chelerythrine, or MCPG, showing that NO is generated by TMT and that associated changes in NOS are regulated by a PKC-mediated mechanism. Oxidative damage by TMT was demonstrated by detection of elevated malondialdehyde levels. It was concluded that low concentrations of TMT (0.01-0.1 microM) cause apoptotic cell death in which oxidative signaling is an important event. Higher concentrations of TMT initiate necrotic death, which involves both an oxidative and a non-oxidative component. TMT-induced necrosis but not apoptosis in granule cells is mediated by glutamate receptors.

Topics: Alkaloids; Animals; Apoptosis; Benzophenanthridines; Catalase; Cells, Cultured; Cerebellum; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; In Situ Nick-End Labeling; L-Lactate Dehydrogenase; Necrosis; Neurons; NG-Nitroarginine Methyl Ester; Nitrites; Phenanthridines; Protein Kinase C; Rats; Reactive Oxygen Species; Receptors, Glutamate; Tetradecanoylphorbol Acetate; Thiobarbituric Acid Reactive Substances; Trimethyltin Compounds

1. Magnesium (Mg)-deficient rats develop a mechanical hyperalgesia which is reversed by a N-Methyl-D-Aspartate (NMDA) receptor antagonist. Given that functioning of this receptor-channel is modulated by Mg, we wondered whether facilitated activation of NMDA receptors in Mg deficiency state may in turn trigger a cascade of specific intracellular events present in persistent pain. Hence, we tested several antagonists of NMDA and non-NMDA receptors as well as compounds interfering with the functioning of intracellular second messengers for effects on hyperalgesia in Mg-deficient rats. 2. Hyperalgesic Mg-deficient rats were administered intrathecally (10 microl) or intraperitoneally with different antagonists. After drug injection, pain sensitivity was evaluated by assessing the vocalization threshold in response to a mechanical stimulus (paw pressure test) over 2 h. 3. Intrathecal administration of MgSO4 (1.6, 3.2, 4.8, 6.6 micromol) as well as NMDA receptor antagonists such as MK-801 (0.6, 6.0, 60 nmol), AP-5 (10.2, 40.6, 162.3 nmol) and DCKA (0.97, 9.7, 97 nmol) dose-dependently reversed the hyperalgesia. Chelerythrine chloride, a protein kinase C (PKC) inhibitor (1, 10.4, 104.2 nmol) and 7-NI, a specific nitric oxide (NO) synthase inhibitor (37.5, 75, 150 micromol x kg(-1), i.p.) induced an anti-hyperalgesic effect in a dose-dependent manner. SR-140333 (0.15, 1.5, 15 nmol) and SR-48968 (0.17, 1.7, 17 nmol), antagonists of neurokinin receptors, produced a significant, but moderate, increase in vocalization threshold. 4. These results demonstrate that Mg-deficiency induces a sensitization of nociceptive pathways in the spinal cord which involves NMDA and non-NMDA receptors. Furthermore, the data is consistent with an active role of PKC, NO and, to a lesser extent substance P in the intracellular mechanisms leading to hyperalgesia.

Topics: 2-Amino-5-phosphonovalerate; Alkaloids; Analgesics; Animals; Benzophenanthridines; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hyperalgesia; Indazoles; Injections, Spinal; Kynurenic Acid; Magnesium Sulfate; Male; Neurons; Nitric Oxide Synthase; Pain Measurement; Phenanthridines; Protein Kinase C; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Spine