Stress-induced depression of motor activity correlates with regional changes in brain norepinephrine but not in dopamine

This experiment examined how inescapable tail shock alters the level of dopamine and norepinephrine within various brain regions of the rat and the relationship of these changes to the depression of motor activity produced by the shock. Following exposure to tail shock that is known to interfere with acquisition of active behavioral tasks, animals were briefly tested for spontaneous motor activity and then sacrificed for neurochemical measures. Norepinephrine and dopamine levels in the frontal cortex, brain stem, striatum, olfactory tubercle, hypothalamus, hippocampus, septum, and amygdala were measured by a sensitive radicenzymatic technique. Exposure to 45 min of tail shock did not alter motor activity significantly, but shock sessions of 60 and 75 min duration produced a marked decrease in motor activity. Levels of dopamine were found to be very little changed in all brain regions studied except for the hypothalamus, in which a substantial rise in dopamine level was observed. Norepinephrine levels, in contrast, fell in many brain regions in response to shock. The fall in norepinephrine levels observed in twi brain regions was significantly correlated with the decline in motor activity (brain stemr=+0.70, hypothalamusr=+0.60) These data suggest that deficits in active motor behavior produced by shock parameters similar to those used in this study may reflect concomitant disturbances of noradrenergic function in specific brain regions.     

Effects of chronic metrifonate treatment on cholinergic enzymes and the blood-brain barrier

After an acute (4 h) treatment with an irreversible cholinesterase inhibitor organophosphate, metrifonate (100 mg/kg i.p.), the activities of both acetyl- and butyrylcholinesterase were inhibited (66.0-70.7% of the control level) in the rat brain cortex and hippocampus. There were no significant changes in the acetyl- and butyrylcholinesterase activities in the olfactory bulb, or in the choline acetyltransferase activity in all three brain areas. After chronic (2 or 5 week) metrifonate treatment (100 mg/kg daily i.p.), the activities of both cholinesterases were substantially inhibited in the rat brain cortex and hippocampus (15.8-31.8% of the control levels), but there was no inhibition of the choline acetyltransferase activity. Moreover, chronic metrifonate treatment did not have any effect on the distribution of the acetylcholinesterase molecular forms. In vitro, metrifonate proved to be a more potent inhibitor of butyryl- than of acetylcholinesterase in both the cortex and the hippocampus. In the hippocampus, the butyrylcholinesterase activity was twice as sensitive to metrifonate inhibition as that in the cortex (IC50 values 0.22 and 0.46 microM, respectively). The effects of chronic (5 week) metrifonate treatment on the blood-brain barrier of the adult rat were examined. The damage to the blood-brain barrier was judged by the extravasation of Evans' blue dye in three brain regions: the cerebral cortex, the hippocampus, and the striatum. No extravasation of Evans' blue dye was found in the brain by fluorometric quantitation. These data indicate that chronic metrifonate treatment may increase the extracellular acetylcholine level via cholinesterase inhibition, but it does not have any effects on the blood-brain barrier. Therefore, it appears reasonable to hypothesize that cholinesterase activities do not play a role in the blood-brain barrier permeability.     

Activity of Na, K-ATPase and the enzymes of intermediate metabolism in the brains of rats exposed to electroshock]

Activity of Na, K -ATPase, acetylcholinesterase (AChE) and glutamic acid decarboxylase (GAD) in the fractions of the rat brain and spinal cord tissue were studied in rats during a single electroshock (ES) and 5 and 30 minutes after it. GAD activity of the synaptosome fraction was shown to decrease insignificantly, but activity of AChE, Na, K -ATPase and possibly of proteolytic enzymes increased 5 minutes after electroshock and became normal in 30 minutes. It is supposed that the revealed inhibition of Na, K -ATPase activity in the "synaptosomes" of the rat brain cortex could be of pathogenetic significance in the origination of the convulsive process.     

Modulation in Acetylcholinesterase of Rat Brain by Pyrethroids In Vivo and an In Vitro Kinetic Study

Abstract: The modulation in acetylcholinesterase (AChE) of rat brain by two pyrethroids—permethrin (PM) and cypermethrin (CPM)—was studied both in vivo and in vitro. PM inhibited AChE activity in all regions of the rat brain (cerebral cortex, cerebellum, corpora striata, brainstem, hippocampus, and hypothalamus) at 4, 8, and 12 h after gastric intubation, whereas CPM elevated the enzyme activity in vivo. Substrate-dependent enzyme kinetic studies have shown that PM and CPM behave as mixed-type inhibitors, as evidenced by alterations in both Michaelis-Menten constant ( K _m) and maximal velocity ( V _max) values. This indicates that both PM and CPM and substrate acetylcholine interact at hydrophobic subsites and may be able to bind simultaneously to the enzyme.     

Effect of chronic administration of propranolol on acetylcholinesterase and 5-hydroxytryptamine levels in rat brain and heart

The effect of chronic administration of propranolol on the rat brain and heart acetylcholinesterase was studied by administering propranolol (5mg/kg body weight) for 14 days. This treatment was found to substantially inhibit the enzyme activity. Levels of 5-hydroxytryptamine were measured in the brain and heart; in brain the levels of 5-hydroxytryptamine increased with propranolol administration, while in the heart there was no change. Effect of different concentrations of propranolol on acetylcholinesterase activity was also studied in vitro and a decreased activity of the enzyme was found in brain and heart homogenates. The significance of these results is discussed in terms of the therapeautic effects of the drug in the control of hypertension.     

Effect of 6-hydroxydopamine on brain and blood catecholamine, ammonia, and amino acid metabolism in rats subjected to high pressure oxygen induced convulsions

Effects of 6-hydroxydopamine (6-OHDA) on rat brain and blood adrenaline (A), noradrenaline (NA), ammonia (NH3), gamma-aminobutyric acid (GABA), and amino acid metabolism prior to and after high pressure oxygen (OHP) induced convulsions have been studied. 6-OHDA reduces GABA and glutamate (Glu) rior to OHP exposure in rat brain so that the concentration is even equal to that seen in nondrugged animals after convulsion. Concomitantly, 6-OHDA reduces the latency of OHP-induced convulsion significantly, and increases brain NH3, glutamine, and asparagine significantly. Although 6-OHDA, in increasing dosage, elevates blood A concentration, convulsion produces a significant further increase in A. Blood NA was not significantly changed in drugged, convulsed animals and was much less than blood NA concentrations in nondrugged convulsed animals. Increasing doses of 6-OHDA also increase NH3 in the blood significantly and convulsion increases its concentration further. Latency of convulsion seems to be related to certain monoamine levels since in some drugged animals where A and total catecholamines are still reduced 96 h after the first of two doses of 6-OHDA, NA concentrations are recovered to relatively normal and the convulsion latency time is also increased although it remains significantly abbreviated from undrugged animals' convulsion time. Low brain GABA levels seem to be a prime effector of convulsive activity.     

Alterations in some membrane properties in rat brain following exposure to lead

The effect of lead exposure on intracellular calcium levels, membrane fluidity, lipid peroxidation, acetylcholinesterase and monoamine oxidase activity and its accumulation in different regions of the brain were studied to understand the molecular mechanism of lead induced neurotoxicity. Lead treatment (20 mg/kg lead nitrate, intraperitoneally, once daily for 15 days) resulted in a significant accumulation of lead in all brain regions with the maximum being in the hippocampus. Levels of glutathione, lipid peroxidation, intracellular calcium and membrane fluidity, as well as the activity of the membrane bound enzymes, acetylcholinesterase and monoamine oxidase, increased to a significant level in certain areas of the rat brain. The results suggest that lead exerts neurotoxic effects by altering certain membrane bound enzymes and may cause oxidative stress.     

Molecular forms of sucrose extractable and particulate acetylcholinesterase in the developing and adult rat brain

The activity of acetylcholinesterase in the rat striatum increased considerably during development, while activities in the cerebellum and midbrain increased only slightly. During maturation the activity of butyrylcholinesterase increased in all the brain regions examined except in cerebellum. The percentage of acetyl-cholinesterase extractable by isotonic sucrose solution from mature striatum was much smaller than those obtained for other regions of the rat brain. For the developing striatum, the percentage of isotonic sucrose extractable activity was almost three times that for adult striatum. Density gradient centrifugation showed that the membrane-bound particulate fraction of adult rat brain was mostly composed of the 10 S form of acetylcholinesterase with little activity of 4 S form of the enzyme. However, a much higher proportion of the 4 S form was found in the isotonic sucrose soluble fraction. In contrast to the particulate fraction from adult brain, that from 6-day old rats contained a much higher proportion of the 4 S form of the enzyme. The sucrose soluble fraction from 6-day old rat brains contained in general much smaller proportion of 4 S form as compared to those from adult rat brains.     

Intracellular distribution of molecular forms of acetylcholinesterase in rat brain and changes after diisopropylfluorophosphate treatment

The subcellular distribution of acetylcholinesterase activities was studied in the striatum and cerebellum of rat brain. The highest percentage of the enzyme activity was found in the crude synaptosomal (P₂) fraction, with striatum much higher than cerebellum. On sucrose density gradient centrifugation analyses all the particulate fractions (P₁, P₂, and P₃) showed a major peak of the 10 S form of acetylcholinesterase activity with very little activity of the 4 S form of the enzyme. The 10 S/4 S ratio was much higher in striatum than in cerebellum. In the soluble fraction (100,000g supernatant) the 10 S form was less than the 4 S form in the adult rat brain, but this was reversed in the 6-day-old rat brain. After diisopropylfluorophosphate administration the recovery of acetylcholinesterase molecular forms in various subcellular fractions differed at different recovery periods. These results indicate that the distribution of molecular forms of acetylcholinesterase in rat brain differs in various subcellular fractions, and also the pattern of distribution differs in different regions of the brain as well as in adult and developing brains.     

Acetylcholine Content in Rat Brain Is Elevated by Status Epilepticus Induced by Lithium and Pilocarpine

The effects of status epilepticus on the concentration, synthesis, release, and subcellular localization of acetylcholine, the concentration of choline, and the activity of acetylcholinesterase in rat brain regions were studied. Generalized convulsive status epilepticus was induced by the administration of pilocarpine to lithium-treated rats. The concentration of acetylcholine in the cortex, hippocampus, and striatum decreased prior to the onset of spike activity or status epilepticus. Once status epilepticus began, the concentration of acetylcholine increased over time in the cortex and hippocampus, reaching peak levels that were 461% and 304% of control levels, respectively, after 2 h of seizures. Such high in vivo levels of acetylcholine had not been reported previously following any treatment. During status epilepticus, the concentration of acetylcholine in the striatum returned to control levels after the initial depression, but did not accumulate to high levels as it did in the other two regions. The in vivo cortical efflux of acetylcholine was also increased during the seizures. Choline levels were increased by status epilepticus in all three brain regions. Inhibition of seizures by pretreatment with atropine blocked the increases of acetylcholine and choline. Synaptosomes prepared from the cortex and from the hippocampus of rats with status epilepticus had elevated concentrations of acetylcholine: in the hippocampus the acetylcholine was principally in the cytoplasmic fraction, whereas in the cortex the acetylcholine was elevated in both the cytoplasmic and the vesicular fractions. The extra acetylcholine was in a releasable compartment, since increased K+ in the media or ouabain increased the release of acetylcholine from cortical slices to a greater extent in tissue from seized rats than from controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of ethanol on methanol-induced oxidative stress and neurobehavioral deficits

The present study examines the efficacy of ethanol as an antidote in methanol neurotoxicity in terms of its effect on antioxidant defense system and behavior. It was observed that acute methanol exposure (7.5 g/kg body weight) led to an increase in lipid peroxidation in various regions of brain. Ethanol administration (7.5 g/kg body weight), on the other hand, was found to accentuate methanol-induced lipid peroxidation. Glutathione levels in brain were significantly reduced in methanol-exposed animals. However, in the coexposed animals, the levels of glutathione were comparable to those observed in controls. The activities of superoxide dismutase and catalase were decreased in the brain following methanol exposure, whereas in methanol- and ethanol-coexposed animals there was no significant effect on these enzymes as compared to methanol-exposed animals. The activity of acetylcholinesterase was significantly reduced in the methanol-exposed animals. On the other hand, acetylcholinesterase activity was not affected in the coexposed animals in comparison to methanol-treated group. Neurobehavioral studies revealed impaired motor and cognitive functions following methanol exposure. In contrast, ethanol exposure ameliorated the behavioral deficits induced by methanol. The findings from the present study suggest the beneficial effect of ethanol on neurobehavioral deficits induced by methanol along with intensification of methanol-induced oxidative stress.     

ANISOMYCIN,ACETOXYCYCLOHEXIMIDE, CYCLOHEXIMIDE,AND PUROMYCIN AS INHIBITORS OF RAT BRAIN ACETYLCHOLINESTERASE IN VITRO1

A kinetic analysis of the interaction of anisomycin, acetoxycycloheximide, cycloheximide, and puromycin with acetylcholinesterase (acetylcholine acetyl-hydrolase, EC 3.1.1.7) in rat brain homogenate shows that all of these protein synthesis inhibitors are also inhibitors or this enzyme. Puromycitl aminonucleoside, a puromycin analog without antibiotic activity, was also found to be an inhibitor of acetylcholinesterase activity much like puromycin. Anisomycin appeared to be a competitive inhibitor whereas all of the other compounds showed mixed inhibition. The apparent 10.5 values for inhibition of rat brain acetylcholinesterase at 50 μM substrate were: anisomycin, 3 mM; acetoxycycloheximide, 1 mM; cycloheximide, 2.2 mM; puromycin, 0.5 mM and puromycin aminonucleoside, 0.6 mM.     

A comparison of the localization of acetylcholinesterase in the rat brain as demonstrated by enzyme histochemistry and immunohistochemistry

The localization of acetylcholinesterase (AChE) as revealed either by enzyme-histochemical or by immunohistochemical methods was compared in distinct regions of the rat brain. In general, the localization of AChE observed was nearly the same, whether revealed by histochemical demonstration of its catalytic activity or by immunohistochemical detection of the enzyme molecule itself, in all regions investigated. Penetration problems of the antibodies, however, arose on strong myelin sheaths of the facial nerve, for instance, where no immunohistochemical staining was found though there was a relatively strong histochemical reaction. These problems could be partly solved by increasing the normal concentration of Triton X-100 added to the immunohistochemical solutions (0.1%) to 2.5%. Furthermore, it seems that sites containing low amounts of AChE could be better detected by the enzyme-histochemical method, whereas the depiction of structures (particularly of nerve fibres) was somewhat sharper with the immunohistochemical method.     

Anticonvulsant Activity of Muscimol and γ-Aminobutyric Acid Against Pyridoxal Phosphate-Induced Epileptic Seizures

The intracerebroventricular injection of pyridoxal phosphate (PLP, 0.125-1.25 μmol/rat) causes epileptic seizures (4 min → 1 min) that are preventable or reversible by GABA (1 μmol/rat), by muscimol (O.025 μmol/rat), or by diazepam (1.75 μmol/rat). At the peak of PLP-induced convulsions, the activities of GAD and GABA-T in 14 regions of rat brain remained unaltered, whereas the concentrations of PLP remained elevated. The PLP-induced convulsion was blocked by DABA (10 μmol/rat) but was not altered by β-alanine (50 μmol/rat). The previous in vitro studies have shown that PLP increases the uptake of [³H]GABA into synaptosomes and inhibits the binding of [³H]GABA to synaptic membranes. These data suggest that PLP-induced convulsion is due to reduced availability of GABA to its recognition sites, rather than to alteration in the activity of GABA metabolizing enzymes, or unavailability of PLP as a coenzyme for GAD and GABA-T. Since the duration of PLP-induced epileptic seizures is short and can be prevented by GABA agonists, PLP may be used as a tool to study the nature of GABA-mediated neuroinhibition and the properties of GABA receptor sites.     

PROPERTIES AND REGIONAL DISTRIBUTION OF TRYPTOPHAN HYDROXYLASE IN THE CHICKEN BRAIN

Tryptophan hydroxylase in young chicken brain had a pH optimum of 7.5–8, depending on the buffer used. It had apparent K_m values for tryptophan and tetrahydrobiopterin of 49 μM and 32 μM respectively. The enzyme in chicken brain, but not rat brain, was cold-shock labile but was stable for up to 4 days at — 20°C. Lability was observed both in tissues and homogenates of these tissues subjected to cold shock, but the extent of loss of activity varied between brain regions. Supernatant fractions did not lose activity after cold shock. The highest level of tryptophan hydroxylase was found in the rostral region of the chicken brainstem. High levels were also found in the caudal region of the brainstem, the midbrain, thalamus, caudate and cerebral cortex. The cerebellum and optic chiasma contained only traces of activity.     

Distributions of choline acetyltransferase and acetylcholinesterase activities in layers of rat superior colliculus

Choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activities were assayed in samples dissected from sagittal sections through rat superior colliculus. The magnitude of ChAT activity was about half to equal that found in rat whole brain in all layers except stratum griseum intermediale, where the average activity was higher than whole brain. AChE activity was three to four times that found in rat whole brain in superficial layers and about the same as average brain in deeper layers, except in the statum griseum intermediale, where the average activity was about twice whole brain. Rostral-caudal gradients in both ChAT and AChE activities occurred in stratum griseum intermediale, with activities in the caudal region of some animals as high as four times those in the rostral. ChAT activity in samples associated with locations of patches or spots of AChE staining product in stratum griseum intermediale was significantly higher than in samples from "nonpatch" regions. Results are discussed relative to inputs into the colliculus, whose terminations may correlate in location with the distributions of the enzyme activities.     

Effect of imipramine hydrochloride on the activity of gamma-aminobutyric acid transaminase in regions of the rat brain

The effect of intraperitoneal injection of imipramine hydrochloride on the activity of gamma-aminobutyric acid transaminase was determined in three regions of the rat brain.The cerebral hemispheres did not show a significant change in the activity of gamma-aminobutyric acid transaminase. Cerebellum and brain stem, both, however, showed a very significant decrease in the activity of the enzyme at 15 and 30 minutes after drug administration. At 90 minutes after drug administration, the activity of gamma-aminobutyric acid transaminase had returned to nearly control values.     

Carbaryl-induced elevation of corticosterone level and cholinergic mechanism

Administration of a single dose (200 mg/kg, p.o.) of carbaryl to rats produced a significant rise in adrenal and plasma corticosterone levels and an increase of tyrosine alpha-ketoglutarate transaminase activity in the liver cytosol. Synaptosomal acetylcholinesterase activity of the hypothalamic and the striatal regions of rat brain was decreased by carbaryl treatment under similar conditions. Pretreatment (0.5 h) with atropine sulphate (10 mg/kg, i.p.) failed to counteract the carbaryl-induced elevation of adrenal and plasma corticosterone levels and hence the liver tyrosine alpha-ketoglutarate transaminase activity. Present results suggest that the carbaryl-induced rise in the corticosterone level in the adrenal gland and plasma is not due to a cholinergic mechanism.     

Transcription-Dependent and -Independent Induction of Cerebral Ornithine Decarboxylase

Ornithine decarboxylase (ODC; EC 4.1.1.17) is a highly inducible, rate-limiting enzyme of the polyamine pathway. We have studied the mechanisms that lead to the induction of ODC activity in response to electrical stimulation in three brain regions. Hippocampal ODC activity was found to exhibit much larger elevations than that of the neocortex and the cerebellum. The levels of ODC gene expression were also followed to examine its relationship to the existing regional differences in ODC activity. In the neocortex, there was an elevation of both the ODC mRNA and enzyme activity. However, the hippocampal ODC mRNA level was not increased by electroconvulsive shock. Furthermore, the effects of hormonal changes and seizures on these regional differences in ODC induction were also examined. Adrenalectomy did not affect ODC activity, but pretreatment with the anticonvulsant MK-801 caused a depression of the induced levels of enzyme activity. Our data suggest that ODC activity in all the brain regions studied is directly elevated by electrically stimulated seizures. However, this induced ODC activity may or may not involve enhanced gene expression.