MECHANISM OF ACTION OF NOTEXIN AND NOTECHIS 11-5 ON SYNAPTOSOMES

The neurochemical activity of notexin and notechis II-5 was investigated using a synaptosomal preparation of rat cerebral cortices. In preparations preincubated with [³H]choline in order to label acetylcholine, the toxins caused a rapid release of the transmitter which was calcium-dependent. The toxins were also potent inhibitors of high affinity choline uptake. Both agents produced a marked depolarization of the synaptosomal preparation as measured by a fluorescent dye and at high concentrations lysed the preparation. At a concentration of 0.1 μM, notexin and notechis II-5 caused a 50% increase in the efflux of lactate dehydrogenase activity. These results, together with electron microscopic observations, indicated that the toxins disrupt the synaptosomal membranes presumably by their inherent phospholipase activity. The release of acetylcholine and inhibition of choline uptake, together with the depolarization of synaptosomal membranes noted in this study, could explain the observed electrophysiological effects of these toxins.     

Lysine production from hydrocarbon by Micrococcus varians 2Fa

A bacterium isolated from Assam (India) soil was found to accumulale l-lysine from hydrocarbon and was identified as a strain of Micrococcus varians. The strain is able to grow and accumulate lysine in a purely synthetic medium though supplementation of the synthetic medium with casamino acids significantly improves the yield. The yield of l-lysine under optimal conditions was found to be 2.6 g X 1(-1) of the compound isolated in crystalline form.     

Effect of selective muscarinic receptor agonists and antagonists on active-avoidance learning acquisition in rats

Effect of some selective muscarinic receptor agonists and antagonists was investigated on learning acquisition in an active-avoidance paradigm in rats which records an anticipatory conditioned avoidance apart from the classical conditioned avoidance response. The muscarinic M1 agonists, arecholine, pilocarpine and McN-A-343, facilitated learning acquisition, which was attenuated by the selective M1 antagonist, pirenzepine. On the other hand, M2 receptor agonist, carbachol, and physostigmine, induced a dose-related dual response, with lower doses retarding and higher doses facilitating the learning acquisition. The former effect was attenuated by gallamine, a muscarinic M2 antagonist, while the latter response was inhibited by pirenzepine, indicating that these putative M2 receptor agonist lose their receptor specificity on dose increment. The selective M2 receptor antagonists, gallamine and AF-DX 116, facilitated learning acquisition, which was inhibited by pirenzepine and the acetylcholine synthesis inhibitor hemicholinium. The results support the cholinergic hypothesis of learning and memory and indicate that M1 receptor agonists and M2 receptor antagonists are likely to prove beneficial in memory deficits. The data also indicates that the clinical dose of some drugs, like physostigmine, needs to be carefully established for optimum therapeutic benefit.     

Effects of muscarinic receptor agonists and antagonists on response of non-extensor rats to maximal electroshock

Rats which do not respond consistently to maximal electroshock by exhibiting the classical hindlimb extensor response, are designated as 'flexors', and can serve as a useful experimental model for investigating seizure mechanisms. 20-25% Charles Foster rats exhibit the flexor status and were used in this study. The flexor rats were converted to extensors by acetylcholine (icv), physostigmine (ip) and the selective muscarinic M1 receptor agonists, arecholine (ip) and McN-A-343 (icv). This conversion of flexors to extensors was significantly attenuated by M1 receptor antagonists scopolamine (ip) and pirenzepine (icv). The M2 receptor agonist, carbachol (icv), had no effect in lower doses but induced conversion of flexor rats to the extensor status only in very high doses which may be due to loss of receptor specificity on dose increment. The M2 receptor antagonists, gallamine (icv) and AF-DX 116 (ip), also induced significant conversion of flexors to extensors, which was dependent upon the availability of neuronal acetylcholine since the effects were attenuated following pretreatment with hemicholinium, an inhibitor of acetylcholine synthesis. The results suggest that the central cholinergic system has a facilitatory pro-convulsant effect, mediated through the muscarinic M1 receptors, an action modulated by the M2 receptors.     

Effects of Hypoxia on the Activity of the Dopaminergic Neuron System in the Rat Striatum as Studied by In Vivo Brain Microdialysis

The purpose of the present study is to clarify the effects of hypoxia on the activity of the dopaminergic neurons in the brain and its mechanism of action. For this purpose, the effects of hypoxia on the extracellular levels of 3,4-dihy-droxyphenylethylamine (dopamine) were examined in the rat Striatum using in vivo brain microdialysis in the presence or absence of pretreatment with either tetrodotoxin (a blocker of voltage-dependent sodium channels) or nomifensine (a blocker of dopamine reuptake). Exposure to various degrees of hypoxia (15, 10, and 8% O₂ in N₂) increased dopamine levels in striatal dialysates to 200, 400, and 1,100%, respectively, of the control value. On reoxygenation, dopamine levels in the dialysates rapidly returned to the control level. Reexposure to hypoxia increased the dopamine levels to the same extent as during the first exposure. After addition of tetrodotoxin (40 mUM) to the perfusion fluid or pretreatment with nomifensine (100 mg/kg, i.p.), exposure to hypoxia no longer increased the dopamine levels. These results suggest that although hypoxia induces an increase in the extracellular dopamine levels (hence, an apparent increase in the activity of the dopaminergic neurons), this increase is not the result of an increase in dopamine release itself, but rather the result of inhibition of the dopamine reuptake mechanism.     

Receptor-operated calcium-channels in isolated rabbit jejunum.

Calcium channels were studied in isolated spontaneously rhythmic rabbit jejunum using the muscarinic agonist carbachol as stimulant. Carbachol failed to produce the characteristic phasic and tonic components of smooth muscle contractions. A variety of chemically distinct calcium antagonists, viz. bepridil, diltiazem, isradipine (PN 200-110), nifedipine, and verapamil, non-competitively inhibited the contractions. Diltiazem was most potent (-logIC50 = 8.30) and bepridil least potent (-logIC50 = 6.19) in inhibiting the contractions. The findings conclude with the presence of pharmacologically distinct receptor-operated calcium-channels, besides the potential-dependent calcium-channels, in the rabbit jejunum.     

Apoptosis. Biochemical events and relevance to cancer chemotherapy.

Two distinct pathways for cell death exist. Compared to necrotic death, physiological or apoptotic cell death is an active suicidal process that consists of a cascade of well-regulated synthetic events. Participation of specific genes in apoptosis, and its possible molecular regulation, are considered in order to investigate the mechanism of cell death induced by some cancer chemotherapeutic agents.     

A T cell nuclear factor resembling NF-AT binds to an NF-kappa B site and to the conserved lymphokine promoter sequence "cytokine-1".

Nuclear extracts from a nontransformed murine T lymphocyte clone contained two inducible factors that bound to a nuclear factor kappa B (NF-kappa B) site. One factor was NF-kappa B, and the other was differentiated from NF-kappa B by its mobility in the electrophoretic mobility shift assay and its lack of sensitivity to protein kinase C depletion. Competition and methylation interference assays showed that the binding site for the novel factor was limited to nucleotides in the 3' half of the kappa B site. This part of the kappa B site resembled sequences in the binding site for a second inducible nuclear factor of T cells, NF-AT, as well as a conserved sequence found in several lymphokine genes, termed "cytokine-1" (CK-1). Competition and methylation interference analysis showed that both NF-AT and CK-1 sequences bound a factor similar to the novel kappa B-binding factor and that binding involved a four-nucleotide sequence (TTCC) that the kappa B, CK-1, and NF-AT sites have in common. The complexes that form with each site have characteristics of NF-AT: they are induced upon T cell receptor stimulation, are sensitive to protein synthesis inhibitors and cyclosporin A, and are not sensitive to protein kinase C depletion. Thus, a factor or factors similar to NF-AT can bind to three distinct promoter sequences which occur commonly in several T cell activation genes. These results raise the possibility that related factors binding to kappa B, CK-1, and NF-AT sequences could play a role in the coordinate induction of T cell activation genes. In addition, our results suggest that kappa B and CK-1 sites represent potential cyclosporin-sensitive promoter elements by virtue of their ability to bind an NF-AT-like factor.