Increased Hippocampal 5-Lipoxygenase mRNA Content in Melatonin-Deficient, Pinealectomized Rats

Abstract: Tryptophan hydroxylase, the initial and rate-limiting enzyme in the biosynthesis of the neurotransmitter serotonin, is activated by protein kinase A and calcium/calmodulin-dependent protein kinase. One important aspect of the regulation of any enzyme by a phosphorylation-dephosphorylation cascade, and one that is lacking for tryptophan hydroxylase, lies in the identification of its site of phosphorylation by protein kinases. Recombinant forms of brain tryptophan hydroxylase were expressed as glutathione S -transferase fusion proteins and exposed to protein kinase A. This protein kinase phosphorylates and activates full-length tryptophan hydroxylase. The inactive regulatory domain of the enzyme (corresponding to amino acids 1–98) was also phosphorylated by protein kinase A. The catalytic core of the hydroxylase (amino acids 99–444), which expresses high levels of enzyme activity, was neither phosphorylated nor activated by protein kinase A. Conversion of serine-58 to arginine resulted in the expression of a full-length tryptophan hydroxylase mutant that, although remaining catalytically active, was neither phosphorylated nor activated by protein kinase A. These results indicate that the activation of tryptophan hydroxylase by protein kinase A is mediated by the phosphorylation of serine-58 within the regulatory domain of the enzyme.     

Behavioural evidence for simultaneous dual changes of 5-HT receptor subtypes: mode of antidepressant action?

Effects of the classic antidepressant imipramine and of an imipramine-like potential antidepressant dihydroergosine were studied in mice, rats and guinea pigs using behavioural models associated with the activation of 5-HT2 and 5-HT1 receptors respectively. Both drugs given in a single dose inhibited the 5-HT2 mediated behaviour for 24 and 48 h respectively and simultaneously stimulated 5-HT1 mediated behaviour for 6 days. Blockade of 5-HT2 receptors could have reduced their inhibitory influence on 5-HT1 receptors. We propose that the interplay between the two receptor subtypes controls the serotoninergic transmission. This idea throws a new light on the mode of action of antidepressants.     

Sex related differences in the response of mice, rats and cats to administration of picrotoxin

Picrotoxin, 2.5 mg/kg, which was subconvulsive in male rats was 92% convulsive in female rats. Four mg/kg of picrotoxin, a dose which did not produce death in the male rats, was 75% lethal in the female rats. Picrotoxin also produced a significantly greater increase in the frequency of the spinal motoneurons discharge in the female than in male rats (444% of control compared to 222% of control). A similar significant difference to the analogous treatment was obtained in the female and male cats (439% of control compared to 368% of control). To counteract the picrotoxin-induced increased frequency of the spinal motoneurons discharge a double dose of diazepam had to be given to females of both species. A sex related difference in the occurrence of convulsions, latency and death following picrotoxin administration was also present in mice. However, mice responded in an opposite direction to rats and cats. Three mg/kg of picrotoxin was 100% convulsive and 27% lethal in male mice, while only 40% convulsive and 0% lethal in female mice. In male mice treated with a 100% lethal dose of picrotoxin, diazepam, 3.0 mg/kg, did not diminish the occurrence of convulsions but reduced the incidence of death to 70%. In equally treated female mice the same dose of diazepam reduced the occurrence of convulsions from 100 to 70% and the incidence of death to 10%. The existence of sex related differences in the response of mice, rats and cats to administration of picrotoxin might have its origin in the dimorphisms of the GABA system in these animal species.     

Induction of Ornithine Decarboxylase by N-Methyl-D-Aspartate Receptor Activation Is Unrelated to Potentiation of Glutamate Excitotoxicity by Polyamines in Cerebellar Granule Neurons

Abstract: Polyamines positively modulate the activity of the N -methyl- d -aspartate (NMDA)-sensitive glutamate receptors. The concentration of polyamines in the brain increases in certain pathological conditions, such as ischemia and brain trauma, and these compounds have been postulated to play a role in excitotoxic neuronal death. In primary cultures of rat cerebellar granule neurons, exogenous application of the polyamines spermidine and spermine (but not putrescine) potentiated the delayed neurotoxicity elicited by NMDA receptor stimulation with glutamate. Furthermore, both toxic and nontoxic concentrations of glutamate stimulated the activity of ornithine decarboxylase (ODC)—the key regulatory enzyme in polyamine synthesis—and increased the concentration of ODC mRNA in cerebellar granule neurons but not in glial cells. Glutamate-induced ODC activation but not neurotoxicity was blocked by the ODC inhibitor difluoromethylornithine. Thus, high extracellular polyamine concentrations potentiate glutamate-triggered neuronal death, but the glutamate-induced increase in neuronal ODC activity may not play a determinant role in the cascade of intracellular events responsible for delayed excitotoxicity.     

Abusive stimulation of excitatory amino acid receptors: a strategy to limit neurotoxicity

Glutamate is an important excitatory amino acid at many central nervous system synapses. After its release from presynaptic nerve terminals, glutamate transiently binds to specific neuronal membrane receptors, which transduce its signal by the generation of intracellular second messengers before being rapidly cleared from the synapse. However, during ischemia, the glutamate concentration at synapses surrounding the focal lesion can be increased for sustained periods of time, resulting in abusive stimulation of glutamate receptors that can eventually be neurotoxic. To develop drugs capable of selectively blocking the pathological effects of glutamate in neurons surrounding ischemic lesions while leaving the physiological actions of glutamate in nonlesioned areas of the brain unaffected, it is essential to delineate glutamate-induced intracellular events that are specific to receptor abuse. This article describes the intracellular sequelae of physiological and pathological glutamate receptor activation and suggests potential targets for such receptor abuse-dependent antagonists (RADAs).     

Putative role of neuronal 5-lipoxygenase in an aging brain.

Aging is associated with increased incidence and/or severity of neurodegenerative pathologies. Oxygen-mediated events are being considered as possible mechanisms responsible for the increasing neuronal vulnerability. Lipoxygenases are enzymes that, as cyclooxygenases (COX), can insert oxygen into the molecule of arachidonic acid and thereby synthesize inflammatory eicosanoids: leukotrienes [due to 5-lipoxygenase (5-LOX) activity] and prostaglandins (via COX activity). It appears that 5-LOX is expressed in central nervous system neurons and may participate in neurodegeneration. 5-LOX-triggered cell death may be initiated by the enzymatic activity of 5-LOX but could also occur via the nonenzymatic actions of the 5-LOX protein; new data point to the possibility that 5-LOX protein exerts actions such as interaction with tyrosine kinase receptors, cytoskeletal proteins, and the nucleus. The expression of neuronal 5-LOX is susceptible to hormonal regulation, presumably due to the presence of hormone-responsive elements in the structure of the 5-LOX gene promoter. The expression of the 5-LOX gene and the activity of the 5-LOX pathway are increased in elderly subjects. One possible mechanism of such 5-LOX up-regulation implies the contribution of aging-associated hormonal changes: relative melatonin deficiency and/or hyperglucocorticoidemia. Thus, the 5-LOX pathway could become a promising target of neuroprotective therapies for the aging brain.     

Glutamate Neurotoxicity Is Independent of Calpain I Inhibition in Primary Cultures of Cerebellar Granule Cells

Glutamate-induced neurotoxicity and calpain activity were studied in primary cultures of rat cerebellar granule neurons and glial cells. Calpain activation, as monitored by quantitative immunoblotting of spectrin, required micromolar concentrations of Ca2+ in neuronal homogenates (calpain I) and millimolar Ca2+ concentrations in glial homogenates (calpain II). Glutamate-induced toxicity and calpain activation were observed in neuronal, but not in glial, cultures. In neurons, calpain I activation by glutamate was dose-dependent and persisted after withdrawal of neurotoxic doses of glutamate. Natural (GM1) and semisynthetic (LIGA4) gangliosides or the glutamate receptor blocker MK-801 prevented calpain I activation and delayed neuronal death elicited by glutamate. GM1 and LIGA4 had no effect on calpain I activity in neuronal homogenates, however. Furthermore, two calpain I inhibitors (leupeptin and N-acetyl-Leu-Leu-norleucinal) prevented glutamate-induced spectrin degradation, but failed to affect glutamate neurotoxicity. These results thus suggest that glutamate-induced neurotoxicity is independent of calpain I activation.