The central mechanism of the immunomodulating action of vasopressin and its structural analogs]

It is shown that the effect of neuropeptides depends on the dose and their structure. Considerable immunostimulation is observed after administration of [8-arginine]vasopressin and des-9-glucine-[8-arginine]vasopressin as against the control. This stimulation is prevented by the preliminary heloperidol blockade of dopaminergic receptors, i.e. it is dopamine-dependent. At the same time, injection of tetrapeptide vesopressin-(2-5) induces deep immunosuppression in all the applied doses. The peptide immunosuppression is shown to be a result of interaction with another neuromediator (serotoninergic); it has no effect under preliminary cuproheptadine blockade of C-2 serotonin postsynaptic receptors. The immunomodulating effect of the neuropeptides studied is central one and is realized via the hypothalamus-hypophysis complex.     

Serotonin reuptake blocker fluoxetine suppresses hippocampal theta oscillation in rabbits during EEG

In earlier studies it has been shown that stimulation of the median raphe nucleus (MR) in awake rabbits decreases the expression and frequency of oscillatory theta activity in the septohippocampal system, and the functional blockade of this nucleus evokes the regular and high-frequency theta rhythm. The present work was aimed at elucidation of serotoninergic influence of MR (which also contains cells of other chemical nature) to the septohippocampal system of theta activity. Serotonin reuptake blocker fluoxetine that increases brain serotonin level was applied. Hippocampal electroencephalogram was recorded in awake rabbits. Bilateral intracerebroventricular infusion of fluoxetine hydrochloride (Sigma, St. Louis; 15 micrograms in 5 microliters saline) in all cases reduced the rhythmic theta activity. In 15 of 18 (83.3%) of experiments the decrease in hippocampal theta oscillations was more than 50% of the control level. The theta band of the spectral density histogram decreased in the mean by 56 +/- 5.8% of the control level (from 10 to 93% in different experiments, p < 0.001). The mean latency of these changes was 3.5 +/- 0.11 minutes (2.9-4.1 min), the effect duration was 64 +/- 3.2 min (45.3-90 min). The mean frequency of the theta waves did not change as compared to the baseline and was equal to 5.25 +/- 0.5 Hz (4.5-6.5 Hz). The fluoxetine-induced reduction of the theta rhythm expression in hippocampus is the evidence of its inhibitory control by serotoninergic brain system. It is suggested that the increase of the frequency of hippocampal theta rhythm after the functional blockade of MR observed in our earlier experiments was the result of a release of the septohippocampal system from the influence of nonserotoninergic neurons (via glutamatergic reticular formation) and/or temporary cessation of the MR interaction with noradrenergic, dopaminergic and glutamate/aspartate systems.     

Central dopaminergic and serotoninergic systems in the regulation of adrenal tyrosine hydroxylase.

Administration of the dopamine receptor agonists apomorphine, piribedil and bromocryptine caused an increase in adrenal tyrosine hydroxylase (TH; tyrosine-3-monooxygenase, EC 1.14.16.2) which could be partially abolished by prior injection of the dopamine blocker haloperidol. Injection of L-dihydroxyphenylalanine, along with the decarboxylase inhibitor carbidopa, also led to a highly significant increase in adrenal TH activity. Intraventricular injection of 5,7-dihydroxytryptamine (DHT), which destroys serotonin neurons, doubled adrenal TH activity in both normal and hypophysectomized rats. Splanchnicotomy abolished this effect of DHT. The increase in enzyme activity mediated by DHT could be partially prevented by peripheral administration of L-5-hydroxytryptophan together with carbidopa. Blockade of serotoninergic functions with the antagonist methiothepin also increased adrenal TH activity. The interrelationship between the dopamine and the presumed serotonin system was investigated. Intraventricular injection of 6-hydroxydopamine partially prevented the DHT-induced increase in adrenal TH activity. Administration of haloperidol to DHT-treated rats had the same effect. This suggests that an intact dopaminergic system is required. When DHT and either apomorphine or piribedil were adminstered simultancously the dopamine agonist-induced increase was potentiated. An intact serotoninergic system is therefore not required for dopamine function. Thus, the increase in adrenal TH activity is associated with either stimulation of central dopamine receptors or destruction of serotonin neurons. It is suggested that dopaminergic and serotoninergic systems are involved in the regulation of adrenal TH and that these systems have net excitatory and inhibitory roles, respectively. Furthermore, the present evidence favors the view that the interaction between the two systems is sequential, with the serotonin system preceding the dopamine one.     

Time Course Changes in the Dopaminergic Nigrostriatal System Following Transection of the Medial Forebrain Bundle: Detection of Oxidatively Modified Proteins in Substantia Nigra

Abstract: We studied the time course of oxidatively modified proteins in the nigrostriatal dopaminergic system following transection of the medial forebrain bundle by quantifying the number of carbonyl groups coupled to striatal and nigral protein homogenates, an index of metal-catalyzed oxidations. We found a striking effect of axotomy on the number of oxidatively modified proteins in the substantia nigra but not in the striatum within the first 5 days postlesion. This effect was correlated with the neurochemical activity of the dopaminergic and serotoninergic systems in the substantia nigra, which suggests a role of dopamine- and serotonin-derived radical oxygen species in the oxidative stress detected in this brain area. We then searched for the type of cell death in the substantia nigra following axotomy. The fragmentation pattern obtained by agarose gel electrophoresis of DNA isolated from nigral tissue was indicative of cell death being entirely necrotic. In fact, no evidence of apoptosis was detected at any postlesion time as revealed by TdT-mediated dUTP-biotin nick end-labeling (TUNEL) staining. The course of necrotic cell death in the substantia nigra coincided with the maximal levels of oxidatively modified proteins in the substantia nigra, suggesting a link between oxidative stress and nerve cell death and also coinciding with the neurochemical activity of both dopaminergic and serotoninergic systems.     

Study of the stimulating effect of the sympathetic stem on the stomach contraction

Stimulation of the sympathetic stem chest portion induces enhancement rather than suppression of gastric contractions. The activating effect is more obvious under conditions of the alpha- and beta-adrenoreceptors blockade with phenolamine and obsidan, and eliminated with the smooth muscle serotoninoreceptors blocking agent lisergol. The findings suggest that the sympathetic stem includes some serotoninergic neural fibres exerting a strong effect upon gastric and intestinal contractions. A previously unknown serotoninergic part of the vegetative nervous system controlling the internal organs functions, seems to exist in the organism.     

Interaction between the serotoninergic and dopaminergic systems in d-fenfluramine-induced activation of c-fos and jun B genes in rat striatal neurons

To test for the relative contributions of the dopaminergic and serotoninergic systems in the striatum to the effects of d-fenfluramine, an indirect serotonin receptor agonist, we assessed the expression of Fos/Jun proteins induced by d-fenfluramine given alone or in the presence of dopaminergic or serotoninergic agents. To determine the neuronal targets of d-fenfluramine in the striatum, we identified the phenotypes of striatal neurons in which d-fenfluramine induced Fos expression. Our results demonstrated that d-fenfluramine evokes nuclear expression of Fos/Jun B proteins in the striatum, and that the Fos expression was dose-dependent and accompanied by transient induction of c-fos mRNA. Fos expression was blocked by p-chloroamphetamine, a serotoninergic neurotoxin. Pretreatment with SCH 23390, a D1-dopamine receptor antagonist, led to a marked decrease in Fos/Jun B expression in the caudoputamen, but not in the cortex, whereas pretreatment with methiothepin, a nonselective serotonin 5-HT1 receptor antagonist, blocked Fos expression completely in the cortex and only partially in the caudoputamen. The expression of Fos/Jun B in the striatum occurred mainly in dynorphin-containing neurons and in a subpopulation of striatal interneurons that exhibited NADPH-diaphorase activity. Most of the enkephalin-containing neurons of the striatum did not show Fos/Jun B staining. These results suggest that the mechanism by which d-fenfluramine induces c-fos and jun B expression in the rat caudoputamen depends at least in part on activation of the dopaminergic system by serotonin.     

Peripheral deletion after bone marrow transplantation with costimulatory blockade has features of both activation-induced cell death and passive cell death

Two major pathways of death of previously activated T cells have been described: activation-induced cell death can be triggered by restimulating activated T cells with high concentrations of Ag, is Fas-dependent, is not influenced by proteins of the Bcl family, and is blocked by cyclosporin A; in contrast, passive cell death is induced by the withdrawal of growth factors and activation stimuli, is Fas-independent, and is blocked by Bcl family proteins. We examined the role of these two forms of cell death in the peripheral deletion of donor-reactive host T cells after allogeneic bone marrow transplantation and costimulatory blockade with anti-CD154 plus CTLA4Ig in two murine models. The substantial decline in donor-reactive CD4 cells seen in wild-type recipients 1 wk after bone marrow transplantation with costimulatory blockade was largely inhibited in Fas-deficient recipients and in Bcl-x(L)-transgenic recipients. We observed these effects both in a model involving low-dose total body irradiation and a conventional dose of bone marrow, and in a radiation-free regimen using high-dose bone marrow transplantation. Furthermore, cyclosporin A did not completely block the deletion of donor-reactive CD4(+) T cells in recipients of bone marrow transplantation with costimulatory blockade. Thus, the deletion of donor-reactive T cells occurring early after bone marrow transplantation with costimulatory blockade has features of both activation-induced cell death and passive cell death. Furthermore, these in vivo data demonstrate for the first time the significance of in vitro results indicating that proteins of the Bcl family can prevent Fas-mediated apoptosis under certain circumstances.     

Nucleotide requirements for the bone marrow heme oxygenase system

Rat bone marrow microsomal heme oxygenase activity has been studied and optimal condition for the measurement of this activity are described. The activity of bone marrow heme oxygenase was linear with time and protein concentration as measured under these assay conditions. Bilirubin formation by the heme oxygenase complex system was observed with either a NADPH generating system or with NADH as electron donor. The enzyme activity for heme degradation supported by NADH proceeds at a comparable rate to that observed with NADPH as reducing equivalent. It thus appears that this oxidation reaction must be more complex than simply involving NADPH as the sole election donor as has been previously proposed.     

The effect of low frequency peripheral stimulation on the tail-flick reflex and evoked cortical potentials in the rabbit

The effects of low frequency peripheral electrical stimulation of innocuous intensity (LES) on the tail-flick latency and the amplitude of the late component of the cortical potentials evoked by single electrical stimuli (20-40 ms latency) were studied in 59 awake and restrained rabbits. Increase of the tail-flick latency and profound decrease of the amplitude of the late positive component of evoked cortical potentials was observed in 50 animals. Five animals of this group and five animals from a group of 9, resistant to electrostimulation, were tested to the analgesic effects of morphine using the same criteria. Positive effects were found in the former but not in the latter group. Naloxone (0.1 mg/kg) profoundly decreased the effects of LES which suggests that an opioid mechanism is involved in the action of conditioning electrostimulation. Pretreatment of the animals with serotonin depletor p-CPA resulted in partial blockade of the effects of LES. The blockade was removed by administration of serotonin precursor 5-HTP. This is in agreement with the hypothesis that the opioid effects are partially exerted through serotoninergic descending pathways.     

Influence of blockade of the dopaminergic D1/D2 receptors on choice behaviour at different by validity reinforcements in cats

The influence of two selective antagonists of the dopaminergic receptors, raclopride (D2) and SCH 23 390 (D1) on behavior of "impulsive" and "self-controlled" cats was similar directed. A selective blockade of the dopaminergic D1/D2 receptors by use of raclopride and SCH 23 390 changed a behavioural choice strategy of two different by validity and delay reinforcements in impulsive cats, but did not change it substantially in "self-controlled" animals. Increase of doses of raclopride and SCH 23 390 led to decrease of a part of the short-latency and increase of a part of the long-latency reactions (pedal pressings) in a total number of all effective responses. This occurred only in "impulsive" animals, that indicates probably the decrease of impulsive properties of their behaviour. Both compounds did not affect the choice strategy (a low quality immediate vs. a high quality delayed reinforcements) in "self-control" animals, but elicited more errors and inhibitory responses in their behavior. The data obtained are not consistent with the "dopamine" hypothesis, which predicts increase of impulsivity under influence of dopaminergic antagonists. The reasons that might lead to the data obtained are discussed.     

Nature of the target cells in bone marrow B-suppressor action studied on a simplified model of suppression activity registration

The addition of bone marrow cells to spleen cells and lymph node cells stimulates by mitogents, but not to fibroblast-like cells, leads to a significant reduction of DNA synthesis in mixed cultures in vitro. The suppression effects appears only in two days and the suppressor cell activity is the stronger, the intensive is the target cell proliferation. It is shown that intact bone marrow cells can suppress the lipopolysaccharide-activated bone marrow cell proliferation in vitro. A conclusion may be draw that cells of the lymphoid system serve as target cells for the bone marrow suppressor cells, and the role of these lymphoid system cells is to control immunogenesis processes by suppressing the target cell proliferation activity in the bone marrow.     

Lipopolysaccharide Intranigral Injection Induces Inflammatory Reaction and Damage in Nigrostriatal Dopaminergic System

Abstract: The pathogenesis of Parkinson's disease is still poorly understood. To address the hypothesis that immunemediated events, such as microglial activation, may be involved in the dopaminergic neurodegeneration, we have studied the effect that intranigral injection of the immunostimulant lipopolysaccharide has on monoaminergic neurotransmitters in rats. Activation of microglial cells, visualized by immunohistochemistry with a specific monoclonal antibody, was already obvious 2 days after injection. In relation to the biochemical parameters studied, we found a significant decrease of dopamine levels in both the substantia nigra and striatum up to at least 21 days after intranigral injection of lipopolysaccharide. This result was supported by the decrease in tyrosine hydroxylase activity and the loss of tyrosine hydroxylase-positive neuronal bodies, shown by immunohistochemistry. These alterations of the dopaminergic system did not reverse during the interval studied (21 days); conversely, the serotoninergic system suffered only transient damage. In addition, we found that the neurotoxic effect of lipopolysaccharide was not mediated by nitric oxide. Based on our results we suggest that the nigrostriatal dopaminergic system is susceptible to damage by inflammatory events and that these may be implicated in neurodegeneration processes such as Parkinson's disease.     

Neutrophil mobilization from the bone marrow during polymicrobial sepsis is dependent on CXCL12 signaling

Neutrophils are essential for successful host eradication of bacterial pathogens and for survival to polymicrobial sepsis. During inflammation, the bone marrow provides a large reserve of neutrophils that are released into the peripheral circulation where they traverse to sites of infection. Although neutrophils are essential for survival, few studies have investigated the mechanisms responsible for neutrophil mobilization from the bone marrow during polymicrobial sepsis. Using a cecal ligation and puncture model of polymicrobial sepsis, we demonstrated that neutrophil mobilization from the bone marrow is not dependent on TLR4, MyD88, TRIF, IFNARα/β, or CXCR2 pathway signaling during sepsis. In contrast, we observed that bone marrow CXCL12 mRNA abundance and specific CXCL12 levels are sharply reduced, whereas splenic CXCR4 mRNA and cell surface expression are increased during sepsis. Blocking CXCL12 activity significantly reduced blood neutrophilia by inhibiting bone marrow release of granulocytes during sepsis. However, CXCL12 inhibition had no impact on the expansion of bone marrow neutrophil precursors and hematopoietic progenitors. Bone marrow neutrophil retention by CXCL12 blockade prevented blood neutrophilia, inhibited peritoneal neutrophil accumulation, allowed significant peritoneal bacterial invasion, and increased polymicrobial sepsis mortality. We concluded that changes in the pattern of CXCL12 signaling during sepsis are essential for neutrophil bone marrow mobilization and host survival but have little impact on bone marrow granulopoiesis.     

Morphological and functional studies on the serotoninergic innervation of the inferior olive

The inferior olive of the cat has, with fluorescence histochemistry, been shown to receive a rich serotoninergic innervation. The distribution of this innervation agrees with the topography of spinal afferent termination as well as the olivo-cerebellar climbing fiber projection. This indicates that different olivary compartments are under different serotoninergic influence. The serotoninergic innervation of the dorsal accessory nucleus (DAO) of the inferior olive of the rat has been identified with electron microscopic radioautography after labelling with 3H-serotonin. The serotoninergic varicosities contain microcanaliculi, tubular-vesicular organelles and large granular vesicles. Few of the serotoninergic varicosities engage in typical synaptic junctions. However, non-junctional varicosities often display other ultrastructural indications of polarity and directed transmitted release. Electrophysiological results indicate that the harmaline-induced tremor, as well as the tremor component of the "serotonin-syndrome", depends on the serotoninergic innervation of the inferior olive. Thus, the sensitivity of different olivary compartments to the induction of rhythmic, synchronous activity by harmaline parallels the distribution of serotoninergic innervation. Neurotoxic destruction of the serotoninergic innervation leads to decreased sensitivity to harmaline. Further, the serotonin receptor agonist 5-methoxy-N,N-dimethyltryptamine, as well as monoamine oxidase inhibition + L-tryptophan loading, leads to rhythmic mass climbing fiber activity in the cerebellum and whole body tremor. A neuromodulatory effect of serotonin on the olivary action potential mechanisms is proposed.     

The interaction of the serotonin and dopaminergic systems of the brain in the mechanisms of latent inhibition in rats

Decrease of serotonin concentration in the septo-hippocampal region of Wistar male rats was obtained by introduction of 5,7-dihydroxytryptamine in the raphe median nucleus. In seven days, after 20 pre-expositions of the conditioned stimulus (presentation of the experimental chamber) conditioned reaction of passive avoidance was elaborated. In sham-operated control the pre-exposition of the conditioned stimulus caused latent inhibition, determined by several parameters: low level of conditioned reaction reproduction, its prolonged preservation at the formed level and nonsubjection to amnesia. Introduction of haloperidol in a dose of 0.5 mg/kg one hour before learning restored the state of latent inhibition disturbed by switching off of the mesolimbic serotonin system. It is supposed that in the state of pre-exposition, the decrease of attention to nonreinforced stimulus takes place because of the intensification of inhibitory influence of the serotoninergic system and reciprocal decrease of dopaminergic system activity.     

The role of the dopaminergic system and GABA-benzodiazepine- ionophore complex in the regulation of memory retrieval

The spontaneous forgetting model has been used to demonstrate the possibility of the memory forgetten trace extraction under the dopamine reuptake blockade by nomifensine and bupropion, increase of its quantity by amfonelic acid, activation of the postsynaptic dopaminergic receptors by (+)3-PPP, blockade of the latter by (-)3-PPP, and under the blockade of separate links of the GABA-benzodiazepine-ionophore complex by bicuculline, picrotoxin, flumazepil and R015-3505. Effectiveness of the neuropharmacological actions improving the memory forgotten trace retrieval is shown to depend upon the duration of the spontaneous forgetting process. The presynaptic receptors are involved in the retrieval process control--improvement of the conditioned habit performance after forgetting due to the activation of presynaptic dopaminergic receptors by specific agonist (-)3-PPP is clearly correlated with the initial retrieval level. The above facts underlie a hypothesis about the neurochemical forgetting mechanisms.     

Role of MAdCAM-1 and its ligand on the homing of transplanted hematopoietic cells in irradiated mice

We examined the expression of VCAM-1 and MAdCAM-1 after bone marrow transplantation (BMT). We also examined the influence of alpha(4)beta(7) integrin blockade on the homing of cells to the bone marrow and spleen. The expression of VCAM-1 and MAdCAM-1 by endothelial cells in the spleen and bone marrow was examined by immunoelectron microscopy using colloidal gold and was analyzed semiquantitatively. To examine the role of alpha(4)beta(7) integrin in donor cells, a homing assay was conducted following alpha(4)beta(7) integrin blockade in bone marrow-derived hematopoietic cells or spleen colony cells. Immediately after BMT, the expression of VCAM-1 and MAdCAM1 markedly decreased, but expression recovered significantly between 12 and 24 h after BMT. VCAM-1 recovered more acutely than MAdCAM-1 from 12 h onward. In the group transplanted with anti-alpha(4)beta(7) integrin antibody-treated bone marrow cells, the numbers of homing cells in the spleen and bone marrow were significantly decreased in an antibody dose-dependent manner. However, the number of homing cells was not different in either the spleen or bone marrow between anti-alpha(4)beta(7) integrin antibody treated and untreated spleen colony cells. It has been reported that alpha(4)beta(1) integrin and its receptor VCAM-1 play major roles in the homing of hematopoietic cells to bone marrow. Our study indicates the importance of MAdCAM-1 and its ligand, alpha(4)beta(7) integrin, in the homing of bone marrow-derived hematopoietic cells, but not spleen colony-derived cells, to both the spleen and bone marrow.     

Dopaminergic parameters in the striatum and substantia nigra of seven strains of mice: Higher density in striatum of CAST compared to BALB mice

Tyrosine hydroxylase activity was assayed in microdissected substantia nigra and striata from seven strains of mice (BALB, CBA, YBR, WB, IS, MOLG, and CAST). In the substantia nigra where tyrosine hydroxylase activity is thought to be proportional to dopaminergic neuron number, only CBA had a different (lower) enzyme activity compared with BALB. However in the striatum, tyrosine hydroxylase activity was larger for IS, MOLG and CAST compared with BALB. Further investigation of the CAST striatum showed that dopamine content and dopamine uptake activity were also higher in comparison with BALB. All three dopaminergic parameters were larger because of lower protein levels in the CAST striatum. A lower absolute amount of glutamic acid decarboxylase activity in CAST versus BALB striatum was consistent with the possibility of a smaller CAST striatum. In contrast to dopamine, the serotonin content in CAST striatum was reduced in proportion to the decrease in protein content. We suggest that the CAST striatum is smaller than BALB striatum and is innervated by proportionally fewer serotoninergic terminals, but the amount of dopaminergic innervation of the CAST striatum is not altered by the size of the target.     

Serotoninergic control of MSH secretion in the eel. Ultrastructural study after 5-hydroxytryptophan treatment.

Injection of 5-HTP induces a melanodisperson ; MSH cells are markedly stimulated :hormone synthesis (development of Golgi area and endoplasmic reticulum) and release (reduction of secretory granules) are observed. This stimulatory serotoninergic pathway seems antagonistic to the dopaminergic system that inhibits MSH secretion in the eel.     

Reduction of food intake by piribedil in the rat: relation to dopamine receptor stimulation

Piribedil, given either intraperitoneally or intracerebroventricularly to rats trained to eat 4 h a day, induced a dose- and time-related anorexia. In this context it was less potent than either amphetamine or fenfluramine.The anorectic effect of piribedil was selectively antagonized by blockade of dopamine (DA) receptors in the central nervous system but not either inhibition of catecholamine synthesis, blockade of α- or β-adrenoceptors or serotoninergic receptors. Also a blocker of “peripheral” DA receptors failed to antagonize piribedil-induced anorexia.Piribedil, as opposed to amphetamine, failed to increase locomotor activity or to induce stereotyped behaviour at doses lower than that required to cause an approximate 80% reduction of food intake.These findings indicate that stimulation of central DA receptors involved in feeding regulation is responsible for the anorexigenic effect of piribedil. This effect in most instances occurs at dose levels of the compound which fail to induce other central stimulant effects.