Fresh Brain Slices Research Articles

Cyanide-induced neurotoxicity: Mechanisms of attenuation by chlorpromazine

Chlorpromazine (CPZ) is an effective cyanide antidote, with its greatest efficacy displayed when combined with the antidotes, sodium nitrite and sodium thiosulfate. Since the central nervous system is a primary target organ in cyanide toxicity, the objective of the present study was to determine the mechanisms by which CPZ prevents cyanide-induced damage in neural systems. KCN (10 m m) increased cytosolic free calcium in rat pheochromocytoma (PC12) cells as indicated by the fluorescent dye quin 2. This was blocked by addition of CPZ (0.1 m m) to the cells 15 min prior to addition of KCN. Incubation of cells with KCN (0.1 m m) increased the levels of lipid conjugated dienes and this was blocked by addition of CPZ (1 μ m). Peroxidation of brain lipids in mice administered KCN (7–15 mg/kg, sc) was also attenuated by pretreatment with CPZ. Furthermore, production of lipid peroxidation in fresh mouse brain slices, following incubation with 0.1 m m KCN, was blocked by simultaneous addition of CPZ. These observations indicate CPZ prevents cyanide-induced calcium influx and decreases peroxidation of membrane lipids. Thus the antidotal activity of CPZ in cyanide toxicity appears to be related to maintenance of cellular calcium homeostasis and membrane integrity.

High-performance liquid chromatographic analysis of in vitro central neuropeptide processing

Reversed-phase high-performance liquid chromatography (HPLC) was used to study and characterize the in vitro proteolytic processing of β-endorphin by twice-washed membrane homogenates. A high-resolution method, capable of separating over 30 different human β-endorphin-related fragments in a single analysis, was used to study the time course of production of specific, biologically active endorphin fragments by membrane-associated proteases. The results demonstrate that frozen (−37°C), postmortem human and rat brains are viable for processing studies and that metabolism proceeds similarly to that in fresh brain homogenates or slices. Significant differences were noted in the formation rates of putative neuroleptic peptides between sex- and age-matched postmortem brain tissues from controls versus postmortem brain tissues from neuropsychiatric patients or drug-treated animals. These data suggest that using HPLC to characterize neuropeptide processing in human or rat membrane-associated enzyme homogenates is both descriptive and quantitative and offers insight into the central regulation of neuropeptide metabolism.

Adenylate cyclase and the search for new compounds with the clinical profile of lithium

It is possible to evaluate the beta-adrenergic receptor-adenylate cyclase complex in the human periphery by measuring the plasma cyclic AMP rise after adrenergic agonists. A clinical trial of the beta 2 adrenergic agonist salbutamol in depression provided an opportunity to test whether adrenergic receptor subsensitivity does occur during clinical antidepressant treatment. After 1 and 3 weeks of oral salbutamol treatment, depression scores declined significantly in 11 depressed patients, while the plasma cyclic AMP response to i.v. salbutamol declined over 60%. The results support the concept that receptor sensitivity changes occur during human antidepressant therapy. Data are presented that Li, too, markedly reduces activity of beta-adrenergic adenylate cyclase in humans. The effect was evaluated by studying the effect of Li at therapeutic serum concentrations on the plasma cyclic AMP response to subcutaneous epinephrine. The Li effect is specific, since the plasma cyclic AMP response to glucagon is not inhibited. In rat cortical slices Li inhibition of noradrenaline-induced cyclic AMP accumulation is clearly demonstrable only at concentrations close to 2 mM Li. However, fresh human brain slices from edges of surgically-removed tumors show Li inhibition at 1 mM Li concentrations. These results imply that in brain as well as periphery, human noradrenergic adenylate cyclase is inhibited by therapeutic concentrations of Li. Demeclocyclin, a tetracycline-derived antibiotic, was found to inhibit noradrenaline-sensitive adenylate cyclase in rat cortical slices and to inhibit amphetamine-induced hyperactivity in rats in an open field. Clinical trials should search for new compounds with the clinical profile of Li.

Adenylate cyclase and the search for new compounds with the clinical profile of lithium.

It is possible to evaluate the beta-adrenergic receptor-adenylate cyclase complex in the human periphery by measuring the plasma cyclic AMP rise after adrenergic agonists. A clinical trial of the beta 2 adrenergic agonist salbutamol in depression provided an opportunity to test whether adrenergic receptor subsensitivity does occur during clinical antidepressant treatment. After 1 and 3 weeks of oral salbutamol treatment, depression scores declined significantly in 11 depressed patients, while the plasma cyclic AMP response to i.v. salbutamol declined over 60%. The results support the concept that receptor sensitivity changes occur during human antidepressant therapy. Data are presented that Li, too, markedly reduces activity of beta-adrenergic adenylate cyclase in humans. The effect was evaluated by studying the effect of Li at therapeutic serum concentrations on the plasma cyclic AMP response to subcutaneous epinephrine. The Li effect is specific, since the plasma cyclic AMP response to glucagon is not inhibited. In rat cortical slices Li inhibition of noradrenaline-induced cyclic AMP accumulation is clearly demonstrable only at concentrations close to 2 mM Li. However, fresh human brain slices from edges of surgically-removed tumors show Li inhibition at 1 mM Li concentrations. These results imply that in brain as well as periphery, human noradrenergic adenylate cyclase is inhibited by therapeutic concentrations of Li. Demeclocyclin, a tetracycline-derived antibiotic, was found to inhibit noradrenaline-sensitive adenylate cyclase in rat cortical slices and to inhibit amphetamine-induced hyperactivity in rats in an open field. Clinical trials should search for new compounds with the clinical profile of Li.

The differential distribution of label following uptake of 3H-labeled amino acids in the dorsal cochlear nucleus of the cat : An autoradiographic study

Light and electron microscopic autoradiography demonstrated a differential distribution of label in the dorsal cochlear nucleus (DCN) of the cat following the incubation of adjacent fresh brain slices with micromolar amounts of different tritiated amino acids in oxygenated Ringer's bicarbonate solutions. After incubation with GABA, glutamic acid, or glycine, label was found primarily over synaptic terminals. It was most heavily concentrated in the outer molecular layer after GABA uptake, over the inner molecular layer after glutamic acid, and more evenly spread over the molecular layer, fusiform cell layer, and deep DCN after glycine. Labeling with aspartic acid was unimpressive. There was diffuse labeling over all of the DCN after incubation with taurine and alanine and, to a lesser extent, with aspartic acid. After incubation with taurine a few cells in the molecular layer were labeled; following alanine more and different cells (probably glia) were labeled. The distribution of labeled synaptic terminals can be correlated with the differential distribution of different classes of axonal input to the DCN.

Electrophysiological, behavioral, and chemical evidence for a noncholinergic, stereospecific site for nicotine in rat brain.

AbstractPharmacological, electrophysiological, and biochemical studies have been conducted with rats in an attempt to demonstrate that some of the behavioral effects of nicotine may involve non‐cholinergic brain sites. The behavioral response consisted of a prostration‐immobilization syndrome following the injection of 1−10 μg of nicotine into the region of the lateral or third transmitters and their antagonists, and a number of other psychotropic drugs were ineffective in either enhancing or antagonizing the action of nicotine. Associated with the behavioral response were distinct electrophysiological patterns recorded from electrodes implanted in the dorsal hippocampus in freely moving rats. The characteristic electrical changes consisted of shifts in the frequency from 6 to 8 per second to 2 to 4 per second, and shifts in the amplitude from 300 to 400 μV to 100 to 200 μV. A number of derivatives of nicotine and piperidine were synthesized which were found to be antagonistic to the psychopharmacological and electrical effects of nictotine. The natural (−)‐nicotine isomer was found to be at least 100 times as effective as the (+)‐nicotine on the brain. It was not possible to demonstrate 3H nicotine bending to neural membrane preparations; however, the sereospecific nicotine binding was demonstrable in fresh brain slices and to glass filters. The Kb was 2.0 × 10−8M for glass.

Nonclassical respiratory activity of tissue slices

The relationship between respiration rate and ambient oxygen tension has been measured for fresh liver, kidney, heart, and brain slices. Statistical procedures are developed for discriminating between the kinetics predicted by the classical diffusion model and those of the Michaelis-Menten equation suggested in earlier papers. In all tissue slices the Michaelis-Menten equation tended to fit better than the classical model. The differences in fit were most pronounced in liver slices and smallest for kidney and brain slices. The classical model is clearly unsatisfactory for liver and heart tissue. Several explanations are discussed including the hypothesis that a significant part of the oxygen transport is by carrier. The possible nature of such a carrier is considered.

Macroscopic demonstration of infarction in fresh brain slices.

Macroscopic demonstration of infarction in fresh brain slices.