The effect of a decrease in cerebral noradrenaline concentrations on the formation and fixation of temporary connections]

A sharp drop in the noradrenaline content (Na) in the brain of mice does not prevent the elaboration and fixation of relatively simple conditioned reflexes, but considerably disturbs the elaboration of more complex conditioned reflexes with delayed reinforcement. The decrease in the NA content in the brain does not appear to influence directly the process of consolidation of temporary connections; impaired fixation of relatively more complex forms of temporary connections is of a secondary nature and is due to disturbances in their formation.     

Effect of an excess of serotonin in the brain on consolidation of temporary connections]

As a result of intravenous administration of 100 mg/kg of 5-oxytryptophane (5-OTP) to mice, the serotonin content in the brain rises in ten minutes by 35 to 37 per cent, and in an hour, by 58 to 60 per cent, and declines in a day to its initial level. In case of elaboration of passive avoidance reflexes (PAR) an hour after 5-OTP injection, their preservation considerably decreases in a day's time, while 5-OTP administration immediately following PARs elaboration does not affect their preservation. However, 5-OTP injected after PARs elaboration to animals subjected to preliminary cooling, sharply deteriorates their preservation. These facts in conjunction with the previously obtained date attest that excess of serotonin in the brain has a negative effect on the earliest stages of consolidation. When the animals are cooled and the brain temperature is reduced, the consolidation processes are retarded; as a result, the temporary connections prove to be still vulnerable to the action of serotonin excess set up after PAR elaboration.     

Effect of a cyclic analog of enkephalin on learning and memory in the mouse

In experiments on mice, the effect of cyclic analogue of enkephalin (CAE) on the processes of learning and memory was studied in control animals and in animals with changed functional state of monoaminergic brain systems. Administration of the peptide to intact animals significantly accelerated the acquisition of conditioned reflexes of two-way avoidance and did not significantly affect the retention of these reflexes and their subsequent reproduction. Retention and reproduction of conditioned reflexes elaborated in one combination, was disturbed. Administration of iprazid did not eliminate the "accelerating effect" of CAE on the formation of conditioned reflexes of the two-way avoidance but sharply disturbed their retention. In such conditions, the amnesing iprazid effect increased still more. Besides, under CAE effect, the activity of acetylcholinesterase in the motor and especially in the visual cortex of the mice increased. The obtained data testify to an important role of the monoaminergic and cholinergic brain mechanisms in realization of CAE effects on the processes of learning and memory.     

The effects of different pretreatment conditions and fixation regimes on serotonin immunoreactivity: a quantitative light microscopic study

An investigation was designed to evaluate the effects of three different fixation regimes on the retention of serotonin-like immunoreactivity in rat midbrain tissue sections. The effects of pretreatment with pargyline-HCl and l-tryptophan on the volume fraction of serotonin-like immunoreactive processes were also examined. Rat brain tissue was fixed with 4% paraformaldehyde (Pf), 4% paraformaldehyde-0.2% picric acid-0.05% glutaraldehyde (Pf-Pa-G), or 4% paraformaldehyde-0.2% glutaraldehyde (Pf-G). Tissue was subsequently processed for immunohistochemistry using a modified peroxidase-antiperoxidase technique and quantified at the light microscopic level by point counting. Fixation with Pf resulted in higher volume fraction determinations of axonal serotonin immunoreactivity than did fixation with Pf-Pa-G or Pf-G. These results provide quantitative data which indicate that even low levels of glutaraldehyde in the fixative significantly decrease serotonin immunoreactivity. Pretreatment with pargyline and tryptophan increased the amount of serotonin immunoreactivity in tissue fixed with Pf-G but not in tissue fixed with Pf. Pretreatment with pargyline and tryptophan is thus recommended when using glutaraldehyde in the fixation process to assure adequate serotonin immunoreactivity. Pretreatment in conjunction with glutaraldehyde fixation, however, appears to cause differential increases in serotonin-like immunoreactivity within brain nuclei that may compromise the interpretation of results.     

Effect of scopolamine on formation and fixation of temporary connections in rats with altered brain serotonin levels

The influence of scopolamine on elaboration and maintenance of conditioned reflexes of two-way avoidance was studied in rats under conditions of excess and deficit of serotonin in the brain. Administration of scopolamine to intact rats accelerated conditioning and did not prevent fixation of the reflex. Administration of scopolamine to animals with a lowered level of serotonin in the brain (by means of para-chlorophenylalanine) impaired conditioning and induced amnesia. In animals with ablated raphe nuclei, the same dose of scopolamine did not prevent elaboration and maintenance of conditioned reflexes. Accumulation of scopolamine in the brain by means of 5-oxytryptophan abolished acceleration of conditioning, which is specific for scopolamine, and affected the preservation of the reflexes. Against the background of the action of iprozid, scopolamine impaired the conditioning and tended to deteriorate the maintenance of conditioned reflexes. It is assumed that the serotoninergic system exerts a modulating influence on the activity of the brain cholinergic and cholinoreactive mechanisms.     

Footshock-induced changes in brain catecholamines and indoleamines are not mediated by CRF or ACTH

Stressful treatments have long been associated with increased activity of brain catecholaminergic and serotonergic neurons. An intracerebroventricular (icv) injection of the corticotropin-releasing factor (CRF) also activates brain catecholaminergic neurons. Because brain CRF-containing neurons appear to be activated during stress, it is possible that CRF mediates the catecholaminergic activation. This hypothesis has been tested by assessing the responses in brain catecholamines and indoleamines to footshock in mice pretreated icv with a CRF receptor antagonist, and in mice lacking the gene for CRF (CRFko mice). Consistent with earlier results, icv administration of CRF increased catabolites of dopamine and norepinephrine, but failed to alter tryptophan concentrations or serotonin catabolism. A brief period of footshock increased plasma corticosterone and the concentrations of tryptophan and the catabolites of dopamine, norepinephrine and serotonin in several brain regions. Mice injected icv with 25 microg alpha-helical CRF(9-41) prior to footshock had neurochemical responses that were indistinguishable from controls injected with vehicle, while the increase in plasma corticosterone was slightly attenuated in some experiments. CRFko mice exhibited neurochemical responses to footshock that were indistinguishable from wild-type mice. However, whereas wild-type mice showed the expected increase in plasma corticosterone, there was no such increase in CRFko mice. Similarly, hypophysectomized mice also showed normal neurochemical responses to footshock, but no increase in plasma corticosterone. Hypophysectomy itself elevated brain tryptophan and catecholamine and serotonin metabolism. Treatment with ACTH icv or peripherally failed to induce any changes in cerebral catecholamines and indoleamines. These results suggest that CRF and its receptors, and ACTH and other pituitary hormones, are not involved in the catecholamine and serotonin responses to a brief period of footshock.     

Effect of carbidine on the dopamine and serotonin receptors of the central nervous system

It has been shown in experiments on mice and rats that unlike haloperidol and sulpiride, carbidine does not influence the intensity and even increases the duration of apomorphine stereotypy. However, similarly to haloperidol carbidine decreases head twitches in mice, induced by administration of 5-hydroxytryptophan. In radioligand-binding experiments in vitro and in vivo carbidine displaced 3H-spiperone but in the brain cortex without having any effect on the binding in the striatum. Carbidine did not affect the dopamine-sensitive adenylate cyclase in rat striatum. Based on these data it is suggested that in contrast to haloperidol and sulpiride, carbidine selectively inhibits serotonin receptors of the brain.     

Tetanus toxin-induced protein kinase C activation and elevated serotonin levels in the perinatal rat brain

A single intracerebral injection of tetanus toxin (TeTox) is able to produce a time-dependent translocation of Ca2(+)-phosphatidylserine-dependent protein kinase C (PKC) in close-to-term rat brain. TeTox-triggered translocation of PKC is dose- and time-dependent, can be prevented by tetanus antitoxin, and does not occur upon administration of toxin fragments B and C. TeTox-triggered PKC translocation is accompanied by a time-dependent increase in brain serotonin (5-HT). Increase of brain 5-HT is independent of monoamine oxidase inhibition by pargyline. Phorbol ester and TeTox cause a significant increase in serotonin while H-7, a kinase inhibitor, does not affect serotonin levels but abolishes the effect of TeTox. Gangliosides prevent TeTox-triggered 5-HT increase. The data are consistent with the possibility that TeTox acts effectively on the serotonergic innervation, presumably in conjunction with PKC to cause accumulation of serotonin.     

Several mechanisms of acetylcholine participation in the processes of formation and fixation of temporary connections

Microphoretic application of acetylcholine (ACH) to the neurones of the rabbit sensorimotor cortex elicits changes of spontaneous and evoked activity which do not correlate with one another and which persist up to two minutes after the end of ACH application. Following the formation of a defensive conditioned reflex to sound, the reactions to ACH in units involved in the formation of the temporary connection are intensified as compared with their reactions before conditioning and with reactions of those neurones which did not elaborate a temporary connection. The possible mechanisms of ACH participation in the processes of elaboration and fixation of temporary connections are discussed.     

Serotonin Depletion-Induced Maladaptive Aggression Requires the Presence of Androgens

The sex hormone testosterone and the neurotransmitter serotonin exert opposite effects on several aspects of behavior including territorial aggression. It is however not settled if testosterone exerts its pro-aggressive effects by reducing serotonin transmission and/or if the anti-aggressive effect of serotonin requires the presence of the androgen. Using the resident intruder test, we now show that administration of the serotonin synthesis inhibitor para-chlorophenylalanine (300 mg/kg x 3 days) increases the total time of attack as well as the percentage amount of social behavior spent on attack but not that spent on threat – i.e. that it induces a pattern of unrestricted, maladaptive aggression – in gonadectomized C57Bl/6 male mice receiving testosterone replacement; in contrast, it failed to reinstate aggression in those not given testosterone. Whereas these results suggest the pro-aggressive effect of testosterone to be independent of serotonin, and not caused by an inhibition of serotonergic activity, the pCPA-induced induction of maladaptive aggression appears to require the presence of the hormone. In line with these findings, pCPA enhanced the total time of attack as well the relative time spent on attacks but not threats also in wild-type gonadally intact male C57Bl/6 mice, but failed to reinstate aggression in mice rendered hypo-aggressive by early knock-out of androgen receptors in the brain (AR^NesDel mice). We conclude that androgenic deficiency does not dampen aggression by unleashing an anti-aggressive serotonergic influence; instead serotonin seems to modulate aggressive behavior by exerting a parallel-coupled inhibitory role on androgen-driven aggression, which is irrelevant in the absence of the hormone, and the arresting of which leads to enhanced maladaptive aggression.     

Differential effects of amylin on memory processing using peripheral and central routes of administration.

Amylin is a peptide hormone secreted from the beta cells of the pancreatic islets. Amylin was administered peripherally or centrally following weak or strong training on footshock avoidance conditioning in a T-maze. Under conditions of weak training, amylin improved memory retention in a dose-dependent manner. Under conditions of strong training, it impaired retention over the same dose range. Central administration of amylin in mice given strong training impaired retention but had no effect on the retention of mice given weak training. These findings suggest that the mechanisms of action by which amylin altered memory processing are different for peripheral and central administration. Peripherally secreted amylin may play a role in the amnesia seen in diabetes and the memory enhancement following glucose administration.     

Lethal synergy between sarcoma-180/TG cells and vesicular stomatitis virus in mice.

An interaction between sarcoma-180/TG cells and vesicular stomatitis virus in adult mice resulted in the rapid onset of extensive mortality. This interaction, termed lethal synergy, occurred only at early stages of ascites induction in animals with no prior virus contact. A significant sparing effect conferred by the serotonin antagonist dibenamine was reversed by the administration of serotonin. The cause of death was not determined, but a mechanism involving hypersensitivity is indicated.     

Tryptophan and serotonin turnover rate in the brain of genetically hyperammonemic mice

An investigation was made into the effects of hyperammonemia on the metabolism of brain serotonin (5-HT). The animal model used was the sparse fur (spf) mouse, which possesses an inborn error of the urea cycle, i.e. an abnormal form of ornithine transcarbamylase. Several indoles were measured in brain and plasma using liquid chromatography with electrochemical detection coupled to an u.v. detection (LCEC-u.v.). In the mutant mice, plasma total tryptophan (TRP) was higher when compared with the controls, while plasma free-TRP portion was unchanged. In these animals, brain TRP was increased whilst the 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) levels were significantly higher in the hypothalamus and midbrain. Experiments with NSD-1015 (100 mg/kg i.p.) indicated that the 5-hydroxytryptophan (5-HTP) synthesis rate was increased in the hyperammonemic mice. Pargyline experiments (100 mg/kg i.p.) confirmed the enhanced brain 5-HT turnover rate in the spf mice. In addition, these experiments led to the conclusion that hyperammonemia does not affect the various rate constants. After administration of NSD-1015, TRP level slightly increased in the spf mouse brains, while it was stationary in those of the controls. This result could indicate an increased activity of hepatic TRP-pyrrolase in the hyperammonemic mice. Valine (VAL) administration (200 mg/kg i.p.) reduced brain TRP content in the two kinds of mice, but its effect was of shorter duration in the spf when compared with the control. Comparison of brain tryptamine level indicated a slight but not significant increase in the mutant mice. The data reported here indicate that hyperammonemia may affect peripheral TRP metabolism with consequences upon brain 5-HT synthesis, which could promote certain neurologic disorders.     

Influence of dieldrin on serotonin turnover and 5-hydroxyindole acetic acid efflux in mouse brain

Chronic administration of dieldrin failed to produce any alteration of brain serotonin, norepinephrine or dopamine in mice, but caused an increase in 5-hydroxyindole acetic acid levels. The turnover rate of serotonin was unaffected by dieldrin. The probenecid induced accumulation rate of 5-hydroxyindole acetic acid was considerably lowered in dieldrin-treated mice. The results suggested a possible influence of dieldrin on 5-hydroxyindole acetic acid efflux from mouse brain.     

Establishment of a Murine Model for Metastasis of Cytokine-Producing Tumor to the Brain

The A375 cell line, derived from human malignant melanoma, has characteristics of interleukin-6 (IL-6) production. By using this cell line, we have investigated a murine metastasis model of IL-6-producing tumors to the brain by injecting A375 cells directly into the left cardiac ventricle. Nude mice were anesthetized with intraperitoneal injection of pentobarbital sodium. Next, A375 cells suspended in phosphate-buffered saline (PBS) were injected into the left cardiac ventricle of mice. An intracardiac injection of 10⁵ cells developed tumor colonies in the brain after 4 to 6 weeks. Metastatic cells were found in every lobe of the brain. An immunocytochemical study revealed IL-6 production by A375 cells at the metastatic sites in the brain. By the transfection of genes encoding proteins into A375 cells, a novel model of protein expression in the brain in vivo could be constructed. Our system does not require great skill. Our experimental model will facilitate future studies of the local effects of proteins in the brain.     

Antidepressant Treatment Outcome Depends on the Quality of the Living Environment: A Pre-Clinical Investigation in Mice

Antidepressants represent the standard treatment for major depression. However, their efficacy is variable and incomplete. A growing number of studies suggest that the environment plays a major role in determining the efficacy of these drugs, specifically of selective serotonin reuptake inhibitors (SSRI). A recent hypothesis posits that the increase in serotonin levels induced by SSRI may not affect mood per se, but enhances neural plasticity and, consequently, renders the individual more susceptible to the influence of the environment. Thus, SSRI administration in a favorable environment would lead to a reduction of symptoms, while in a stressful environment might lead to a worse prognosis. To test this hypothesis, we treated C57BL/6 adult male mice with chronic fluoxetine while exposing them to either (i) an enriched environment, after exposure to a chronic stress period aimed at inducing a depression-like phenotype, or (ii) a stressful environment. Anhedonia, brain BDNF and circulating corticosterone levels, considered endophenotypes of depression, were investigated. Mice treated with fluoxetine in an enriched condition improved their depression-like phenotype compared to controls, displaying higher saccharin preference, higher brain BDNF levels and reduced corticosterone levels. By contrast, when chronic fluoxetine administration occurred in a stressful condition, mice showed a more distinct worsening of the depression-like profile, displaying a faster decrease of saccharin preference, lower brain BDNF levels and increased corticosterone levels. Our findings suggest that the effect of SSRI on depression-like phenotypes in mice is not determined by the drug per se but is induced by the drug and driven by the environment. These findings may be helpful to explain variable effects of SSRI found in clinical practice and to device strategies aimed at enhancing their efficacy by means of controlling environmental conditions.     

Brain region-specific alterations of 5-HT2A and 5-HT2C receptors in serotonin transporter knockout mice

The aim of the present studies was to determine the effects of reduced or absent serotonin (5-HT) transporters (5-HTTs) on 5-HT2A and 5-HT2C receptors. The density of 5-HT2C receptors was significantly increased in the amygdala and choroid plexus of 5-HTT knockout mice. On the other hand, the density of 5-HT2A receptors was significantly increased in the hypothalamus and septum, but reduced in the striatum, of 5-HTT knockout mice. However, 5-HT2A mRNA was not changed in any brain region measured. 5-HT2C mRNA was significantly reduced in the choroid plexus and lateral habenula nucleus of these mice. The function of 5-HT2A receptors was evaluated by hormonal responses to (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI). Oxytocin, but not adrenocorticotrophic hormone or corticosterone, responses to DOI were significantly greater in 5-HTT knockout mice. In addition, Gq and G11 proteins were not significantly changed in any brain region measured. The present results suggest that the constitutive alteration in the function of 5-HTTs changes the density of 5-HT2A and 5-HT2C receptors in a brain region-specific manner. These changes may not be mediated by alterations in their gene expression or in the level of Gq/11 proteins. The alterations in these receptors may be related to the altered behaviors of 5-HTT knockout mice.     

Interstrain differences in the serotonin and 5-hydroxyindoleacetic acid levels in the postnatal ontogeny of C57BL/6J and BALB/cLac strain mice

The content of serotonin and 5-hydroxy-indole-acetic acid (5-HIAA) was determined in the brain stem and hemispheres in 1 and 3 months old C57BL/6J and BALB/cLac mice. The characteristic dynamics of serotonin and its metabolite content related to the age was found in different brain regions and proved to be similar in both the strains, but the rate of serotonin system development in C57BL/6J mice was higher than in BALB/c Lac mice. An intensive catabolism of serotonin, possibly, related to the reaction to new environment was noted in the newborn animals. Sex differences in the rate of serotonin system maturation and serotonin and 5-HIAA content were shown for 12--16 days old mice: 12 days old males were characterized by more intensive metabolism than females while 16 days old males had less serotonin than females.     

Improved Production of Human Immunoglobulin γ1 Chain Fc Polypeptides in Escherichia coli and Their Properties

Exposure to some xenobiotics (pentobarbital, 3-terf-butyl-4-methoxyphenol (BHA), chloretone (acetone chloroform), 1, l-bis-(p-chlorophenyl)-2,2,2-trichloroethane (DDT) and polychlorinated biphenyls (PCB)) for a 5 hr period increased the concentrations of brain serotonin and 5-hydroxyindole acetic acid (5HIAA). The decrease in the brain serotonin level elicited by /7-chlorophenylalanine (PCPA), an inhibitor of serotonin synthesis, was prevented by the concomitant administration of chloretone. The administration of both chloretone and pargyline (an inhibitor of monoamine oxidase) caused significant elevation of the brain 5HIAA level as compared with that in a pargyline control, however, the concentration of brain serotonin was not different between pargyline alone and chloretone plus pargyline. These results show that the increase in the brain serotonin level caused by chloretone is not due to acceleration of brain serotonin synthesis, but to retardation of the degradation of brain serotonin, and the increase in brain 5HIAA caused by chloretone may be due to the reduced removal of 5HIAA from the brain. Chloretone plus pargyline caused significant elevation of hypothalamus catecholamines, as compared to in the pargyline control, so the catecholamine turnover rates may be accelerated by the administration of chloretone.     

Formation, fixation and reproduction of temporary connections during adaptation to physical loading

It is shown that long-term adaptation to physical loads of various intensity and duration has an influence on formation, fixation and reproduction of temporary connections in rats. Periodicity of conditioned activity has been found in the process of protracted training. After twenty days of adaptation to submaximal physical loads, the processes of fixation and reproduction of temporary connections improve whereas with elongation of adaptation the conditioned activity somewhat deteriorates. In conditions of long-term training by maximal physical loads, both elaboration and preservation of conditional reflexes in all adaptation terms is affected.