Influence of short-lasting bilateral clamping of carotid arteries (BCCA) on GABA turnover in rat brain structures

We have previously shown that short-lasting reduction of cerebral blood flow by bilateral clamping of carotid arteries (BCCA) results in long-lasting increase in regional GABA concentration and decrease in seizure susceptibility in rats. In the present experiments, the effect of BCCA on GABA turnover and the enzymes involved in GABA synthesis and degradation were studied in rats. Regional GABA turnover was measured by means of GABA accumulation induced by the GABA-transaminase (GABA-T) inhibitor aminooxyacetic acid (AOAA). Fourteen days after BCCA, GABA turnover was significantly increased in hippocampus, substantia nigra and cortex, but not different from sham-operated controls in several other brain regions, including striatum, hypothalamus and cerebellum. The activity of glutamate decarboxylase (GAD) measured ex vivo did not show any changes in investigated structures, while the activity of GABA-T was slightly increased in hippocampus. The increased GABA turnover in some brain regions may explain our previous findings of increased GABA content in these brain regions and decreased sensitivity of BCCA treated animals to the GABA_A-receptor antagonist bicuculline.     

Effects of morphine administration and its withdrawal on rat brain aminopeptidase activities

The endogenous opioid neuropeptide system seems to be involved in the neural processes which underlie drug addiction. Several studies have reported that the administration of morphine induces changes in the levels and/or activity of endogenous opioid peptides (enkephalin, dynorphin) and their precursors in specific brain regions of the adult CNS. The aim of this work was to study the effects of chronic morphine exposure and its withdrawal on certain aminopeptidases capable of degrading opioid peptides in brain areas including the amygdala, hypothalamus, hippocampus, striatum and brain cortices. In animals treated with morphine, aminopeptidase N presented higher enzyme activity levels in the striatum, the hypothalamus and the amygdala compared to control animals, although statistically significant differences were observed only in the case of the striatum. In addition, the activity of soluble puromycin-sensitive aminopeptidase (PSA) was found to be higher in the frontal cortex of these rats. In contrast, rats experiencing withdrawal symptoms presented decreased levels of aminopeptidase activity in certain brain areas. Thus, the activity of aminopeptidase N in the hippocampus and soluble puromycin-sensitive aminopeptidase in the frontal cortex were found to be lower in rats experiencing naloxone precipitated withdrawal symptoms, compared to the corresponding controls. Finally, the activity of the three studied aminopeptidases in vitro was unaltered by incubation with morphine, suggesting that the observed effects are not due to a direct action of this opioid upon the aminopeptidases. The results of the present report indicate that aminopeptidases may play an important role in the processes of tolerance and withdrawal associated with morphine administration.     

Changes in brain serotonin metabolism and [3H]-serotonin receptor binding during recall of conditioned passive avoidance in rats

The content of serotonin and its metabolite 5-hydroxyindoleacetic acid, monoamine oxidase activity, and [3H]-serotonin radioligand receptor binding were examined in the prefrontal cortex, striatum, amygdala, hippocampus and periaqueductal gray matter at different time after one-trial passive avoidance training of rats. Changes in the serotonergic activity were observed only in rats, which showed retrieval of conditioned passive avoidance response. No serotonergic changes were found immediately and one day after training. Also, there were no changes in trained rats without retrieval of conditioned passive avoidance response or rats with experimental amnesia. The pattern of the involvement of brain structures in the retrieval process was also revealed. [3H]-serotonin binding was decreased in the amygdala, periaqueductal gray matter and striatum, whereas it did not change in the prefrontal cortex and hippocampus. At the same time, the serotonin content in these structures did not differ from that of intact rats. Deamination of serotonin by monoamine oxidase and active transport of 5-hydroxyindoleacetic acid from nerve terminals were increased in the amygdala and periaqueductal gray matter, whereas in the striatum serotonin catabolism was decreased. The obtained differences in serotonin catabo- lism suggest that the decrease in receptor binding of serotonin in these brain structures is provided by different synaptic processes: presynaptic changes in the striatum and postsynaptic receptor changes in the amygdala and periaqueductal gray matter. It is concluded that the decrease in the serotonergic activity in the amygdala and periaqueductal gray matter represents one of the mechanisms activating the emotiogenic system mediating the memory trace retrieval in inhibitory avoidance learning.     

In vivo studies of the effect of magnetic field exposure on ontogeny of choline acetyltransferase in the rat brain

Developmental increases of the activity of choline acetyltransferase (ChAT) were examined in the brains of fetuses and offspring from parent rats continuously exposed to a 500 mG, 60 Hz circularly polarized (CP) magnetic field (MF) prior to pregnancy, and further, during pregnancy and lactation. In developing rats between 12 days and 20 days of embryogenesis that were housed in a control unit, i.e., nonexposed to MF, the specific activity of ChAT in whole brain specimens increased from 2.4% to 6.9% of adult activity, while specific activity of ChAT in rat brain specimens between 12 days of embryogenesis and 10 days of postpartum increased from 2.4% to 21.6% of adult activity. On the other hand, the specific activity of ChAT in whole brain specimens from rats under housed MF exposure conditions was found to increase from 2.6% to 6.7% of adult activity between 12 days and 20 days of embryogenesis and from 2.6% to 21.6% of adult activity between 12 days of embryogenesis and 10 days postpartum. Furthermore, the effect of the same test magnetic environment on the specific activity of ChAT in the brains of parental rats was examined in order to determine whether magnetic field exposure of parental rats might reflect onto the development of fetal brain. It was observed that continuous exposure of parental rats to a 500 mG, 60 Hz CP MF did not show any significant changes in the specific activity of ChAT in the septodiagonal band complex, dorsal and ventral hippocampus, striatum, and frontoparietal cerebral cortex, as compared with the same brain regions of control subjects. As far as the development of cholinergic neural circuits was concerned, these test magnetic environments did not interfere in their development and it suggested that parental-fetal intercommunication might provide relatively stable states for neural development, even under these test magnetic environments. © 1993 Wiley-Liss. Inc.     

Corticosterone administration and lipid metabolism in brain regions during development.

Effect of corticosterone on lipid contents of different brain regions and the effect of age on the sensitivity of these regions to corticosterone have been studied. Corticosterone administration (40 mg/kg body wt, sc) to 17-day-old rat for 3 days led to significant decrease in phospholipid content of cerebellum and increase in cholesterol contents of hippocampus and striatum. However, there was no effect on cerebral cortex and brain stem lipids. This alteration in lipids was associated with decrease in [U-14C] glucose incorporation into cholesterol and phospholipids, decrease in plasma beta-hydroxy butyrate levels and increase in beta-hydroxy butyrate dehydrogenase activity in hippocampus and striatum, thereby suggesting that suppression of glucose utilization by corticosterone was compensated by higher utilization of ketone bodies for lipid synthesis in these regions. The sensitivity to corticosterone appears to be age-specific as, at 20-day, cerebellum, hippocampus and striatum were susceptible, at 10-day only hippocampus and at 40- and 90-day none of these regions responded to the treatment.     

Acetylcholinesterase activity in brain slices of rats subjected to cooling

The acetylcholinesterase (AChE) activity is studied in rat slices of the cerebral cortex, corpus striatum, hypothalamus and medulla oblongata of rats during hypothermia (20 degrees C) and also 1 and 7 days after the posthypothermal period. Cooling of animals down to 20 degrees C is accompanied by an increase in the AChE activity in the brain both under incubation temperature of 20 degrees and 37 degrees C. Under prolonged hypothermia the AChE activity in the investigated brain regions, except for corpus striatum, returns to the control level. By the 7th day of posthypothermal period the AChE activity in corpus striatum, hypothalamus and medulla oblongata does not restore completely. The most substantial changes in the AChE activity both under hypothermia and posthypothermal period occur in corpus striatum, which obviously reflects its complicated functional role.     

Age-related and diurnal changes in Met5-Enk-Arg6-Phe7 and Met5-enkephalin contents of pituitary and rat brain structures

The beta-endorphin, met5-enkephalin-arg6-phe7 (MEAP) and met5-enkephalin (ME) changes related to age and diurnal rhythms were studied in various regions of rat brain and in the pituitary by specific radioimmunoassays. The contents of MEAP, met5-enkephalin and beta-endorphin were higher in the pituitary of old rats (18 months old) than that of young rats (23 days old) while the content of these opioid peptides was higher in the hypothalamus of young rats than in that of old rats. Beta-endorphin was also higher in the striatum of 23 days old rats, but no age-associated changes were observed in the hippocampus, brain stem or cortex. In the diurnal rhythm study, it was found that in the hypothalamus and striatum of the adult rat (2-3 months old), both MEAP and ME contents were higher at mid-dark than at mid-light and that in the intermediate posterior lobe of the pituitary, the ME content was also higher at mid-dark.     

Increased analgesic potency of morphine and increased brain opioid binding sites in the rat following chronic naltrexone treatment

Implantation of rats with prolonged-release naltrexone pellets increased both morphine's analgesic potency in the tailflick assay and radiolabeled opioid binding in the brain. The increases in both radiolabeled opioid binding and morphine potency were time-dependent. Implantation for 24 hours did not increase binding, whereas increases of approximately 45% were seen following 8 days of implantation. Similarly, morphine's analgesic potency, measured as ED50 values, was increased by 50% following 8 days of exposure to naltrexone while a 24 hour exposure had no significant effect.     

Food deprivation-induced changes in the level of opioid peptides in the pituitary and brain of rat

Following 1-4 days of food-deprivation (FD) male rats were sacrificed. The pituitary and different regions of brain were analyzed for beta-endorphin-like immunoreactivity (beta-EI), dynorphin (dyn) and methionine-enkephalin (ME) content by RIA. Pituitary beta-EI increased by 16, 28 and 43% on days 2, 3 and 4 of FD. In striatum also, beta-EI increased by 140 and 176% on days 2 and 3 of FD. Dyn level in pituitary was not affected but decreased in hypothalamus by 20% and in striatum by 73% on the 4th day of FD. There was a significant decrease (33-55%) in ME levels in striatum, hippocampus and cortex on 4th day of FD. When food-deprived rats were fed for 24 hr, concentration of most of the opioid peptides returned to basal level. These results suggest that FD in rats affects the opioid peptide levels in a differential manner.     

Ethylene dibromide: effects of paternal exposure on the neurotransmitter enzymes in the developing brain of F1 progeny

The effects of ethylene dibromide (EDB) exposure to male rats on several neurotransmitter enzymes have been examined in various brain regions of the F1 progeny, from 7 to 90 days of age. The choline acetyltransferase activity was significantly increased at 21 days old, in most brain regions studied in the F1 progeny of the EDB-treated males, but not at 7, 14 or 90 days old. The acetylcholinesterase activity was altered in different brain regions of the F1 progeny of the EDB-exposed males at both 14 and 21 days old but not at 7 or 90 days old. Glutamic acid decarboxylase activity was increased in corpus striatum but decreased in frontal cortex only at 21 days of age. These neurochemical changes in the developing brain of F1 progeny of EDB-treated males at low doses may be associated with behavioral abnormalities observed early in their development.     

Lithium suppresses elevated behavioural activity and brain catecholamines in developing hyperthyroid rats.

Neonatal hyperthyroidism in rats induced by daily administration of L-triiodothyronine for 30 days since birth resulted in a significant rise in mobility and the metabolism of brain norepinephrine and dopamine. Whereas administration of lithium carbonate (60 mg/kg ip) to normal rats for 6 days produced no effect on spontaneous locomotor activity and increased the synthesis and possibly release of this monoamine in several brain regions, this antimanic drug antagonized the L-triiodothyronine-stimulated increases in mobility as well as norepinephrine and dopamine metabolism of hypothalamus, midbrain, striatum, and cerebral cortex. Furthermore, lithium treatment restored the activity of catechol-O-methyl transferase (EC 2.1.1.6) in young hyperthyroid rats to virtually normal limits. Our data suggest that antiphasic or damping effects of lithium upon mood swings is controlled, at least in part, by catecholaminergic systems in the brain. The interrelationship between brain catecholamines and thyroid hormones seems to be important to our understanding of the action of lithium in affective illness.     

Protein synthesis rates in rat brain regions and subcellular fractions during aging

In vivo protein synthesis rates in various brain regions (cerebral cortex, cerebellum, hippocampus, hypothalamus, and striatum) of 4-, 12-, and 24-month-old rats were examined after injection of a flooding dose of labeled valine. The incorporation of labeled valine into proteins of mitochondrial, microsomal, and cytosolic fractions from cerebral cortex and cerebellum was also measured. At all ages examined, the incorporation rate was 0.5% per hour in cerebral cortex, cerebellum, hippocampus, and hypothalamus and 0.4% per hour in striatum. Of the subcellular fractions examined, the microsomal proteins were synthesized at the highest rate, followed by cytosolic and mitochondrial proteins. The results obtained indicate that the average synthesis rate of proteins in the various brain regions and subcellular fractions examined is fairly constant and is not significantly altered in the 4 to 24-month period of life of rats.A preliminary report of these results was previously presented at: WFN-ESN Joint Meeting on: Cerebral Metabolism in Aging and Neurological Disorders, Baden, August 28–31, 1986.     

Widespread alterations in central noradrenaline,dopamine, and serotonin systems in the Brattleboro rat not related to the local absence of vasopressin

A comprehensive study of monoamine transmitter and metabolite concentrations measured by HPLC was undertaken in female (vasopressin-deficient) Brattleboro rats as compared to Long Evans rats. Noradrenaline was significantly increased in 8 out of 13 dissected brain regions, whereas concentrations of the metabolite 3-methoxy-4-hydroxyphenylglycol were not altered. The increases were not restricted to areas which are normally innervated by vasopressin-containing neurons. Serotonin was increased in 6 and dopamine in 4 regions and this was accompanied in some areas by increases in the metabolites 5-hydroxyindolacetic acid and dihydroxyphenylacetic acid. Only in the striatum, cerebellum, and the medulla-pons no changes could be detected in any of the compounds of interest. These results show that the long term absence of vasopressin in Brattleboro rats appears to be associated with increases in monoamine transmitter contents and decreased metabolite/transmitter ratios. The regional distribution of these changes does not bear any relationship to the regional distribution of vasopressin cell bodies or nerve endings.     

An analogue of thyrotropin-releasing hormone, DN1417, decreases naloxone binding in the rat brain

We explored whether thyrotropin-releasing hormone may affect opioid receptors in the rat brain. Adult male rats were intraperitoneally injected twice a day with varying doses of DN1417, a potent analogue of thyrotropin-releasing hormone, for 2 days, and opioid receptors of the brain (hypothalamus, striatum, hippocampus, midbrain, ponsmedulla, and cortex) homogenates were determined using [3H]naloxone. Intraperitoneal injection of DN1417 in a dose of 0.3 mg/100 g body wt resulted in a significant reduction in naloxone binding of the striatum as compared with the saline-injected group, whereas naloxone binding of other brain regions was not affected by DN1417. DN1417 produced a dose-dependent decrease in naloxone binding of the striatum. The affinity constant of naloxone binding was similar between the saline- and DN1417-injected groups. In vitro addition of DN1417 did not interfere with the brain naloxone binding. The distribution of 3H-labeled DN1417 injected peritoneally did not differ among the brain regions. The present data imply that the opioid antagonistic action of thyrotropin-releasing hormone may be due, at least in part, to the significant decrease in the striatal opioid binding in the rat brain.     

Enhanced Brain Cell Proliferation Following Early Adrenalectomy in Rats

We have previously demonstrated an increase in adult brain DNA content in rats adrenalectomized on postnatal day 11. The present studies examined cell proliferation in cerebral cortex, cerebellum, hippocampus, and midbrain-diencephalon following adrenalectomy at this age. Compared to sham-operated controls, adrenalectomized animals showed increased [3H]thymidine incorporation into DNA (measured at 1 h following a pulse injection) in all brain regions at 7 and 14 days postsurgery. In some areas, the effect was already present as early as 2 days following adrenalectomy. Chronic replacement with corticosterone prevented this increase in DNA labelling in a dose-dependent manner. When cell proliferation in the cerebral cortex and cerebellum was independently assessed by measuring changes in thymidine kinase activity, enzyme activity was significantly elevated in both areas at 7 and 14 days postsurgery. Finally, histological examination of the cerebellar cortex suggested a delayed disappearance of the external granular layer in several cerebellar lobules of adrenalectomized animals. Overall, these findings indicate that day-11 adrenalectomy leads to a prolonged stimulation of mitotic activity in areas where cell formation at this time is exclusively glial (i.e., cerebral cortex and mid-brain-diencephalon) as well as in areas where postnatal neurogenesis is also occurring (cerebellum and hippocampus). It is hypothesized that this stimulation results from the removal of a tonic inhibitory effect exerted by circulating glucocorticoids in the normal intact animal.     

Differential Effect of Cerebral Ischemia on Monoamine Content of Discrete Brain Regions of the Mongolian Gerbil (Meriones unguiculatus)

The effect of bilateral cerebral ischemia on noradrenaline, dopamine, and serotonin concentrations in six brain regions (i.e., the cerebral cortex, striatum, hippocampus, midbrain-diencephalon, cerebellum, and pons-medulla oblongata) was examined in the gerbil stroke model. The relative changes in regional cerebral blood flow after bilateral common carotid occlusion were also assessed using the radioactive microsphere technique. At 1 h after bilateral carotid occlusion, a significant decrease of monoamine concentration was observed in the cerebral cortex, striatum, hippocampus, and midbrain-diencephalon whereas no significant change was detected in the cerebellum and pons-medulla oblongata. The fall in NA content was most prominent in the cerebral cortex and hippocampus and percentage reductions of dopamine and serotonin were greatest in the striatum and cerebral cortex, respectively. These results suggest that the monoamine neurons in various brain regions might have different vulnerabilities to ischemic insult and show no evidence of transtentorial diaschisis.     

Changes in food intake and glucosensing function of hypothalamus and hindbrain in rainbow trout subjected to hyperglycemic or hypoglycemic conditions

To evaluate the possible role of glucose in the control of food intake (FI) in fish and the involvement of glucosensing system in that role, we have subjected rainbow trout (via intraperitoneal injections) to control, hyperglycemic (500 mg kg(-1) glucose body mass) or hypoglycemic (4 mg kg(-1) bovine insulin) conditions for 10 days. The experimental design was appropriate since hypoglycemia and hyperglycemia were observed the first 5 days after treatment and changes observed in metabolic parameters in liver were similar to those of fish literature. Hyperglycemic conditions elicited small changes in FI accompanied by increased glucose and glycogen levels, glucokinase (GK) activity and glycolytic potential in hypothalamus and hindbrain. In contrast, hypoglycemic conditions elicited a marked increase in FI accompanied by decreased glucose and glycogen levels and GK activity in the same brain regions whereas both regions displayed different responses in glycolytic potential. These results allow us to hypothesize that, despite the relative intolerance to glucose of carnivorous fish, changes in plasma glucose levels in rainbow trout detected by glucosensing areas in brain regions (hypothalamus and hindbrain) are integrated in those or near areas eliciting a response in FI, which was more important under hypoglycemic than under hyperglycemic conditions.     

Alterations in physiological functions and in brain monoamine content in the sympathectomized rats

In the present study, we conducted pre-ganglionic decentralization (or sympathetic trunk resection) of the superior cervical ganglia and observed alterations in several physiological functions and in the monoamine content of different brain regions. Over an ambient temperature range of 8-30 degrees C, these sympathectomized rats maintained their rectal temperatures within a normal limit displayed by the intact controls. These sympathectomized animals, although showing no change in the level of spontaneous pain threshold or motor activity, did display an increased sensitivity of analgesic responses to morphine administration or locomotor stimulant responses to amphetamine administration. Biochemical examination revealed that these sympathectomized animals had a higher level of norepinephrine, dopamine or 5-hydroxytryptamine in the hypothalamus, as well as a higher level of dopamine in the corpus striatum. However, in the brainstem, these sympathectomized animals had a unaltered monoamine level. The data indicate that, in a sympathectomized condition, changes in the monoamine content of different brain regions may be correlated with the above-mentioned alterations in somatosensory and motor neural functions.     

Opioids and efflux transporters. Part 4: Influence of N-substitution on P-glycoprotein substrate activity of noroxymorphone analogues

The efflux transporter protein P-glycoprotein (P-gp) is capable of affecting the central distribution of diverse neurotherapeutics, including opioid analgesics, through their active removal from the brain. P-gp located at the blood brain barrier has been implicated in the development of tolerance to opioids and demonstrated to be up-regulated in rats tolerant to morphine and oxycodone. We have previously examined the influence of hydrogen-bonding oxo-substitutents on the P-gp-mediated efflux of 4,5-epoxymorphinan analgesics, as well as that of N-substituted analogues of meperidine. Structure–activity relationships (SAR) governing N-substituent effects on opioid efficacy is well-established, however the influence of such structural modifications on P-gp-mediated efflux is unknown. Here, we present SAR describing P-gp recognition of a short series of N-modified 4,5-epoxymorphinans. Oxymorphone, naloxone, naltrexone, and nalmexone all failed to demonstrate P-gp substrate activity, indicating these opioid scaffolds contain structural features that preclude recognition by the transporter. These results are examined using mathematical molecular modeling and discussed in comparison to other opioid scaffolds bearing similar N-substituents.     

Distinctions in metabolism of serotonin and dopamine in rat brain during latent inhibition

Latent inhibition (LI) is a behavioral phenomenon, in which repeated presenting of a non-reinforced stimulus retards conditioning to this stimulus when it is coupled with a reinforcer. In order to find specific serotonin (5-HT- and dopamine (DA) changes mediating the LI, the 5-HT and DA metabolism was investigated in certain brain regions. Oxidative deamination of 5-HT and DA by monoamine oxidase (MAO) was determined in the prefrontal cortex, striatim, amygdala, and hippocampus at preexposure and testing stages of the LI using the passive avoidance procedure in rats. Preexposed animals demonstrated high MAO activity for 5-HT deamination in the amygdala and striatum and lower MAO activity for DA deamination in the amygdala and hippocampus. After testing the LI, a high level of 5-HT deamination by MAO was revealed in the amygdala, white the lower level of 5-HT deamination by MAO was shown in the prefrontal cortex. At the same time, no changes in DA metabolism were found in all the brain regions studied. Thus, the role of dopaminergic system in the LI effect may be limited by the preexposure stage. The obtained evidence suggests that the enhanced 5-HT activity in the amygdala and striatum induced by the preexposed stimulus is a principal biochemical mechanism underlying the LI.