Activation of CB1 cannabinoid receptors impairs memory consolidation and hippocampal polysialylated neural cell adhesion molecule expression in contextual fear conditioning.

We investigated the role of CB1 receptors in hippocampal-dependent memory consolidation mediated by polysialylated neural cell adhesion molecule (PSA-NCAM) during contextual fear conditioning (CFC). The CB1 receptor agonist 3-(1,1-dimethylheptyl)-(-)-11-hydroxy-Delta(8)-tetrahydrocannabinol (HU-210) (0.1 mg/kg) was given immediately after training during the memory consolidation phase, and freezing behavior was measured 24 h after conditioning. Administration of HU-210 attenuated freezing behavior measured in CFC. Western blot analysis showed that CFC induced a decrease in the expression of NCAM-180, but did not change the level of NCAM-140 and increased PSA-NCAM expression measured 24 h after training in the rat hippocampus. HU-210 (0.1 mg/kg) injection did not affect the reduction in NCAM-180 levels induced by CFC, but it blocked the increase in PSA-NCAM expression. Since the dentate gyrus (DG) of the hippocampus is known to be involved in memory consolidation and expresses a high level of PSA-NCAM protein, we measured the effects of CFC and HU-210 administration on PSA-NCAM-immunoreactive (IR) cells in the DG. CFC caused an increase in the number of PSA-NCAM-IR cells in the DG, but not K(i)-67- or doublecortin (DCX)-IR cells. This increase in PSA-NCAM-IR cells was abolished by HU-210 injection. Administration of the CB1 receptor antagonist N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM-251) (3 mg/kg immediately before HU-210) inhibited the effects of HU-210 on freezing behavior and PSA-NCAM expression in the DG. These results indicate that activation of CB1 receptors disturbs consolidation of fear memory in CFC, likely by affecting PSA-NCAM expression in the DG, which plays an important role in synaptic rearrangement during the formation of memory traces.

Aryl hydrocarbon receptor-mediated apoptosis of neuronal cells: a possible interaction with estrogen receptor signaling.

Activation of aryl hydrocarbon receptors (AhRs) induces neuronal damage, but the mechanism by which this occurs is largely unknown. This study evaluated the effects of an AhR agonist, beta-naphthoflavone, on apoptotic pathways in mouse primary neuronal cell cultures. beta-Naphthoflavone (0.1-100 micronhanced caspase-3 activity and lactate dehydrogenase (LDH) release in neocortical and hippocampal cells. These data were supported at the cellular level with Hoechst 33342 and calcein AM staining. alpha-Naphthoflavone inhibited the action of beta-naphthoflavone, thus confirming specific activation of AhRs. A high-affinity estrogen receptor (ER) antagonist, ICI 182,780, and a selective estrogen receptor modulator (SERM), tamoxifen, enhanced beta-naphthoflavone-mediated apoptosis. Another SERM, raloxifene, and an ERalpha antagonist, methyl-piperidino-pyrazole, did not affect beta-naphthoflavone-induced caspase-3 activity. However, they inhibited beta-naphthoflavone-induced LDH release at a late hour of treatment, thus suggesting delayed control of AhR-mediated neuronal cell death. The apoptotic effects of beta-naphthoflavone were accompanied by increased levels of AhRs, and these receptors colocalized with ERbeta as demonstrated by confocal microscopy. These data strongly support apoptotic effects of AhR activation in neocortical and hippocampal tissues. Moreover, this study provides evidence for direct interaction of the AhR-mediated apoptotic pathway with estrogen receptor signaling, which provides insight into new strategies to treat or prevent AhR-mediated neurotoxicity.

Triclocarban Disrupts the Epigenetic Status of Neuronal Cells and Induces AHR/CAR-Mediated Apoptosis.

Triclocarban is a phenyl ether that has recently been classified as a contaminant of emerging concern. Evidence shows that triclocarban is present in human tissues, but little is known about the impact of triclocarban on the nervous system, particularly at early developmental stages. This study demonstrated that triclocarban that was used at environmentally relevant concentrations induced apoptosis in mouse embryonic neurons, inhibited sumoylation, and changed the epigenetic status, as evidenced by impaired activities of HDAC, sirtuins, and DNMT, global DNA hypomethylation, and alterations of methylation levels of bax, bcl2, Ahr, and Car genes. The use of selective antagonists and specific siRNAs, which was followed by the co-localization of aryl hydrocarbon receptor (AHR) and constitutive androstane receptor (CAR) in mouse neurons, points to the involvement of AHR and CAR in triclocarban-induced neurotoxicity. A 24-h treatment with triclocarban enhanced protein levels of the receptors which was paralleled by Car hypomethylation and Ahr hypermethylation. Car hypomethylation is in line with global DNA hypomethylation and explains the increased mRNA and protein levels of CAR in response to triclocarban. Ahr hypermethylation could reflect reduced Ahr mRNA expression and corresponds to lowered protein levels after 3- and 6-h exposures to triclocarban that is likely related to proteasomal degradation of activated AHR. We hypothesize that the triclocarban-induced apoptosis in mouse neurons and the disruption of epigenetic status involve both AHR- and CAR-mediated effects, which may substantiate a fetal basis of the adult onset of neurological diseases; however, the expression of the receptors is regulated in different ways.

Impact of postnatal blockade of N-methyl-D-aspartate receptors on rat behavior: a search for a new developmental model of schizophrenia.

The malfunction of glutamatergic neurotransmission in the neonatal or postnatal periods may be a risk factor for the appearance of neuroanatomical, neurochemical or functional changes that are characteristic of schizophrenia. Thus, the present study was undertaken to investigate whether blockade of N-methyl-d-aspartate (NMDA) receptors in the postnatal period influences rat behavior in tests characterizing schizophrenia-like deficits such as psychomotor agitation, impairments of sensorimotor gating, working memory, and intensity of social interactions. (E)-2-amino-4-methyl-5-phosphono-3-pentenoic acid (CGP 40116), a competitive antagonist of NMDA receptors, was given postnatally (1.25 mg/kg on days 1, 3, 6, 9; 2.5 mg/kg on days 12, 15, 18; and finally 5 mg/kg on day 21, all injections s.c.), and rats were tested at 60 days old. We found that blockade of NMDA receptors in the postnatal period led to an enhancement of exploration, mimicking psychomotor agitation, impairments in sensorimotor gating as measured by a prepulse-evoked inhibition of acoustic startle response, and an impaired working memory, as measured by an increase in the latency to achieve accurate rate of response in the delayed alternation task. Decreases in non-aggressive social interactions and increases in aggressive interactions were also observed. In addition to cognitive deficits typical of schizophrenia, rats treated postnatally with NMDA receptor antagonists also showed higher level of fear exhibited in the elevated plus maze. Thus, the blockade of NMDA receptors in the postnatal period may model deficits that are characteristic of schizophrenia.

Kainate receptor agonists, antagonists and allosteric modulators.

Interest in kainate receptors has increased over the past few years. Our understanding of their physiology and pharmacology has improved markedly since their original cloning and expression in the early 1990s. For example, agonist profiles at recombinant kainate receptors have been used to identify and distinguish kainate receptors in neurons. Furthermore, the development of selective antagonists for kainate receptor subtypes has increased our understanding of the functional roles of kainate receptors in neurons and synaptic transmission. In this review we described the activity of agonists and antagonists at kainate receptors and their selectivity profiles at NMDA and non-NMDA receptors.

WAY 100135, an antagonist of 5-HT1A serotonin receptors, attenuates psychotomimetic effects of MK-801.

In the present study, we investigated whether the antagonist of 5-HT1A receptors, WAY 100135, was capable of modifying the psychostimulant and psychotomimetic effects of MK-801, a non-competitive antagonist of NMDA receptors. It was found that: 1) WAY 100135 (10 and 20 mg/kg, but not 1.25, 2.5, and 5 mg/kg) transiently, in a dose dependent manner, attenuated the locomotor stimulant effects of MK-801 (0.4 mg/kg). Given alone, WAY 100135 had no effect on the locomotor activity of rats; 2) WAY 100135 (1.25 and 2.5 mg/kg, but not 10 or 20 mg/kg), attenuated or abolished the disruptive effects of MK-801 on the sensorimotor gating measured in a prepulse-induced inhibition of the acoustic startle response paradigm. WAY 100135 in all tested doses had no effect on the sensorimotor gating or amplitude of the acoustic startle response; 3) WAY 100135 (1.25, 2.5 mg/kg, but not 5 mg/kg) attenuated the detrimental effects of MK-801 on working memory and selective attention, measured in a delayed alternation task. Again, given alone, WAY 100135 did not influence the behavior of rats in that experimental paradigm; and 4) MK-801 (0.4 mg/kg) had no effect on the 5-HT1A receptor mRNA level in rat hippocampus, measured 2 and 24 hours after MK-801 administration. These data indicate that 5-HT1A receptors might be involved in the psychotomimetic effects of non-competitive NMDA receptor antagonists. In addition, 5-HT1A serotonin receptor antagonists and partial agonists may have potential antipsychotic properties.

The corticosterone synthesis inhibitor metyrapone decreases dopamine D1 receptors in the rat brain.

Experiments were performed to examine the effect of metyrapone, an inhibitor of corticosterone synthesis, on the level of dopamine D1 receptors and their transcripts in the caudate-putamen, nucleus accumbens and olfactory tubercle of the rat brain. The binding to dopamine D1 receptors was measured by receptor autoradiography using the specific D1 receptor antagonist [3H]SCH 23390. The level of dopamine D1 receptor messenger RNA was determined by in situ hybridization histochemistry. The results obtained have shown that metyrapone (two injections of 150 and 50 mg/kg, i.p., given 20 and 3 h before killing, respectively) induced a decrease in the D1 receptor-specific binding in the studied areas of the rat brain. In the caudate putamen, the decrease in [3H]SCH 23390 binding was stronger in the medial (31-39%) than in the lateral part (24-27%). Decreases similar to those in the caudate-putamen were observed in the nucleus accumbens (21%) and olfactory tubercle (32%). Furthermore, metyrapone decreased the level of dopamine D1 receptor messenger RNA in the caudate putamen (17-28%), nucleus accumbens (20%) and olfactory tubercle (18%). In conclusion, our study indicates that glucocorticoids might be involved in the regulation of dopamine D1 receptor level in the rat brain. since metyrapone (which inhibits the synthesis of these hormones) decreases the messenger RNA encoding D1 receptor synthesis, as well as the specific binding to this receptor.

Distribution of dopamine D1 receptors in the nucleus paraventricularis of the hypothalamus in rats: an immunohistochemical study.

The present study investigated the distribution of dopamine D1 receptor protein in the nucleus paraventricularis of the hypothalamus. It was found that the nucleus paraventricularis of the hypothalamus contains a relatively large number of cells which are positive for presence of dopamine D1 receptor protein. The vast majority of dopamine D1 receptor-positive neurons was found in the magnocellular part, but they were also present in considerable quantity in the parvocellular part of this subregion of the hypothalamus. When measured by the Western blot technique, the quantity of D1 receptor protein found in the paraventricular nucleus of the hypothalamus was at the level found in the prefrontal cortex. It was also found that dopamine D1 receptor protein was present in neurons constitutively displaying phosphorylated CREB protein, i.e. neurons which are, as might be speculated, under the tonic influence of neurotransmitters whose receptors operate via cAMP and pCREB as second or third messengers. The presence of dopamine D1 receptors in the nucleus paraventricularis of the hypothalamus may suggest, at an anatomical level, that these receptors are involved in controlling the release of hormones, as well as their synthesis at the level of transcription, which is regulated by phosphorylation of CREB protein. Finally, the present immunocytochemical findings offer an anatomical substrate for the role of dopamine and its receptors of D1 subtype in the regulation of the activity of paraventricular neurons seen in the functional studies.

c-Fos proteins, induced by the serotonin receptor agonist DOI, are not expressed in 5-HT2A positive cortical neurons.

In the present study, we tried to find out whether the expression of c-Fos proteins induced by DOI, an agonist of 5-HT2A/2C receptor subtypes is colocalized with 5-HT2A receptor protein in cortical neurons. 5-HT2A receptor protein was found in two major neuronal elements: dendritic processes (seen in layers II/III-V) and less abundantly in cell bodies (layer V). In our experiment, DOI (8 mg/kg) induced a robust appearance of c-Fos proteins mainly in neuronal nuclei of the upper part of layer V/IV, and a moderate amount of sparsely distributed nuclei in deep cortical layers (V and VI). It was found that c-Fos proteins never occurred in cortical neurons, which were immunopositive for the presence of 5-HT2A receptor protein. It is concluded that the induction of c-Fos proteins expression by DOI though initiated by activation of 5-HT2A receptors, requires the involvement of intermediate neurotransmitter(s). Additionally, our study indicates that the appearance of DOI-induced c-Fos proteins cannot be used as a simple and direct marker of localization and site of activation of 5-HT2A receptors.

Single doses of MK-801, a non-competitive antagonist of NMDA receptors, increase the number of 5-HT1A serotonin receptors in the rat brain.

In the present study, we investigated the impact of MK-801, a non-competitive NMDA receptor antagonist, on the density of serotonergic receptors of the 5-HT1A subtype and on the metabolism of serotonin in various regions of the rat brain containing terminals and cell bodies of serotonergic neurons. The binding of [3H]8-OH-DPAT to 5-HT1A serotonin receptors was increased after MK-801 (0.4 mg/kg) as was shown by autoradiographic studies in the frontal, cingulate and part of enthorinal cortex, subregions of the hippocampus and raphe nuclei. The above receptor changes were observed at 2 h and, in some brain regions, at 24 h after MK-801. In saturation binding studies, an increase in the Bmax value in the rat hippocampus was found after MK-801 (0.4 mg/kg) while no changes being noted in the Kd value. MK-801 (0.4 mg/kg) increased the concentration of the serotonin metabolite 5-HIAA in the prefrontal cortex and hippocampus, respectively, at 2 and 3 or 3 h after administration, being without effect on the level of serotonin. In the dorsal raphe nucleus, MK-801 (0.4 mg/kg) decreased the level of serotonin without affecting the level 5-HIAA (0.5 h after administration) or increased the level of 5-HIAA without altering the concentration of serotonin (3 h after administration). It is concluded that single administration of MK-801 may alter the density of serotonergic 5-HT1A receptors and in consequence influence the function of the central nervous system associated with activation of 5-HT1A receptors.

Ipsapirone enhances the dopamine outflow via 5-HT1A receptors in the rat prefrontal cortex.

In the present study, we investigated both the effect of ipsapirone on the dopamine outflow and its selectivity towards 5-HT1A receptors in the rat prefrontal cortex. Using a brain microdialysis method in freely moving animals, it was found that ipsapirone, 5 and 10 mg/kg dose-dependently enhanced the outflow of dopamine, while 2.5 mg/kg was ineffective. The above effects of ipsapirone were mimicked by buspirone (2.5 and 5 mg/kg), another 5-HT1A receptor agonist, but not 1-PP (1-pyrimidinylpiperazine, 5 mg/kg)-a centrally active metabolite of ipsapirone. The effect of ipsapirone (10 mg/kg) on the dopamine outflow in the rat prefrontal cortex was antagonized by 1-(2-methoxyphenyl)-4-[4-(2-phthalimido)butyl]piperazine (NAN-190, 1 mg/kg) and (N-tert-butyl-3-(4-(2-methoxyphenylpiperazin-1-yl)-2- phenylpropionamide (WAY 100135, 10 mg/k.g.), i.e. substances with agonistic/antagonistic and antagonistic properties in relation to 5-HT1A receptors, respectively. NAN-190 (1 mg/kg) enhanced the outflow of dopamine, while WAY 100135 (10 mg/kg) failed to alter it. It is concluded that 5-HT1A receptor agonists may be involved in the regulation of dopaminergic neurotransmission in the rat prefrontal cortex and may have therapeutic potential in the treatment of disorders associated with dysfunction of the mesocortical dopaminergic system.

Imipramine increases the 5-HT1A receptor-mediated inhibition of hippocampal neurons without changing the 5-HT1A receptor binding.

The effect of repeated treatment with imipramine on the 5-HT1A receptor-mediated inhibition of a population spike was studied in the rat CA1 hippocampal region ex vivo. Serotonin (5-hydroxytryptamine, 5-HT) and the selective 5-HT1A receptor agonist 8-hydroxy-2-(dipropylamino)tetralin (8-OH-DPAT) decreased dose-dependently the amplitude of population spikes; this effect was blocked by the selective 5-HT1A receptor antagonist (S)-N-tert-butyl-3-[4-(2-methoxyphenyl)piperazin-1-yl]-2-phenylpro panamide dihydrochloride [(S)-WAY 100135]. Repeated (14 days, twice daily), but not single, administration of imipramine (10 mg/kg) shifted the dose-response curves for serotonin and 8-OH-DPAT to the left. Repeated treatment with imipramine did not change the density of 5-HT1A receptors in the hippocampus as measured by autoradiography using [3H]8-OH-DPAT as a ligand. The latter findings indicate that the imipramine-induced increase in the responsiveness of hippocampal neurons to stimulation of 5-HT1A receptors may not involve an increase in the density of this receptor subtype. To find out whether the efficacy of the postreceptor transduction mechanism is changed by repeated treatment with imipramine, we examined the effect of baclofen. The baclofen-induced inhibition of the population spike was not changed by imipramine. Our results suggest that repeated treatment with imipramine induces sensitization to the inhibitory effects of 5-HT1A receptor agonists in the hippocampus.

The impact of a competitive and a non-competitive NMDA receptor antagonist on dopaminergic neurotransmission in the rat ventral tegmental area and substantia nigra.

The study compares effects of the competitive and non-competitive NMDA receptor antagonists, CGP 40116 and MK-801 respectively, on the metabolism of dopamine and on the density of D-1 and D-2 dopaminergic receptors in the rat ventral tegmental area and substantia nigra. The effects of CGP 40116 were tested in a range of doses which either were devoid of or had locomotor- or stereotypy-stimulating effects. It was found that (1) CGP 40116 given in a dose of 5 mg/kg enhanced the locomotor activity of rats and evoked a stereotypy-like activity; doses of 1.25 and 2.5 mg/kg were devoid of such effects; (2) CGP 40116 (5 mg/kg) enhanced the concentrations of dopamine, DOPAC and HVA in the ventral tegmental area, whereas the lowest dose, 1.25 mg/kg was without effect; a dose of 2.5 mg/kg increased the concentration of dopamine only; the only effect of CGP 40116 (5 mg/kg) observed in substantia nigra, was an increase in dopamine concentration; its doses of 1.25 and 2.5 mg/kg were ineffective. (3) MK-801 (0.2 and 0.4 mg/kg) enhanced the concentrations of dopamine, DOPAC and HVA in both structures. A dose of 0.1 mg/kg increased the dopamine concentration only. The effects of MK-801 in substantia nigra were quantitatively weaker than those observed in ventral tegmental area. (4) Both CGP 40116 (5 mg/kg) and MK-801 (0.4 mg/kg) evoked alterations in the density of dopaminergic receptors. D-2 receptors, were up-regulated by MK-801 in ventral tegmental area and subregions of substantia nigra, i.e. pars compacta and pars reticulata, whereas CGP 40116 evoked similar effects in ventral tegmental area only. D-1 receptors in pars compacta and pars reticulata of substantia nigra were down-regulated after administration of either drug. It is concluded that competitive NMDA receptor antagonists in doses which evoke hyperlocomotion and stereotypy-like activity, may have a substantial impact on the dopaminergic neurotransmission in the rat ventral tegmental area and substantia nigra, similar to that described for MK-801, a non-competitive NMDA receptor antagonist. The obtained results may suggest that CGP 40116 and, possibly, other competitive NMDA antagonists may have dopaminomimetic properties, and that their clinical potentials may be limited by the risk of evoking dopamine-dependent psychotomimetic and abusing effects, similar to those described for MK-801.

Freeze-dried vaccine against Rinderpest: stability and activity study.

A freeze-dried vaccine against Rinderpest was prepared from modified virus multiplied in calf kidney cell culture. Characteristics of the vaccine are as follows: high titre after freeze-drying (10(4) CCID50/dose), well-adapted freeze-drying stabilizer which ensures maintenance of the infective titre of the vaccinal virus, even under severe conditions (3.5 days at +45 degrees C), use of an appropriate solvent: magnesium sulphate molar solution or more simply physiological saline (for stability after reconstitution even at high temperatures--up to 4 h at +45 degrees C). The activity of the vaccine, tested in cattle by antibody titration and resistance to specific challenge perfectly satisfies requirements set by the WHO and OIE.

Homologous protection but lack of heterologous-protection by various species and types of Bartonella in specific pathogen-free cats.

Cat-scratch disease (CSD) is caused by Bartonella henselae, and possibly by B. clarridgeiae. In immuno-compromised persons, B. henselae is one of the agents causing bacillary angiomatosis. Domestic cats are the main reservoir of these bacteria, which are transmitted primarily from cat to cat by fleas. Possible strategies to prevent the spread of infection among cats are to eliminate flea infestation or to prophylactically immunize cats. In order to develop an appropriate vaccine, it is important to determine if cats become resistant to re-infection by the same strain or various types or species of Bartonella. In a series of experiments, 21 SPF cats were experimentally infected by the intradermal route with 10(5)-10(10) colony-forming units/ml of either B. henselae type II (17 cats), or a new strain 'Humboldt' isolated from a mountain lion (4 cats). The cats were bled weekly to every other week for determination of bacteremia and specific antibody production. After they cleared their infection, they were challenged by a homologous or heterologous strain of Bartonella: 10 cats were challenged with B. henselae type II, three cats with B. henselae type I, four cats with B. clarridgeiae and four cats with the 'Humboldt' strain. Seven of these cats received a third inoculum dose resulting in three cats sequentially infected with sequence B. henselae type II/B. henselae type II/'Humboldt', two cats with sequence B. henselae type II/'Humboldt'/B. clarridgeiae, and two cats with the sequence 'Humboldt'/B. henselae type II/'Humboldt'. All cats challenged with a homologous strain remained abacteremic after challenge and had an increased IgG antibody titer. All cats challenged with either a different Bartonella species or type became bacteremic. The few cats receiving a third inoculum with a strain homologous to the initial strain remained abacteremicafter that challenge. All cats infected with B. clarridgeiae suffered relapsing bacteremia compared to only 36% of the B. henselae infected cats and 22% of the 'Humboldt'-infected cats (p=0.008). The duration of bacteremia was significantly longer in B. henselae primary-infected cats (mean: 34 weeks) than B. henselae heterologously challenged cats (mean: 9 weeks) (p=0.014). These data clearly indicate the lack of cross-protection between B. henselae and B. clarridgeiae and furthermore, indicate the lack of protection between B. henselae types I and II, and a wildlife isolate. A vaccine strategy for CSD prevention in domestic cats will require a multivalent vaccine approach.

Cortical localization of dopamine D4 receptors in the rat brain--immunocytochemical study.

Using polyclonal antibody against dopamine D4 receptor we investigated cortical distribution of D4 receptors, with the special emphasis on regions of the prefrontal cortex. Prefrontal cortex is regarded as a target for neuroleptic drugs, and engaged in the regulation of the psychotic effects of various substances used in the experimental modeling of schizophrenia. Western blot analysis performed on samples from the rat cingulate, parietal, piriform cortices and also striatum revealed that antibody recognized one main band of approximately 40 kD, which corresponds to the predicted molecular weight of D4 receptor protein. In immunocytochemical studies we found D4 receptor-positive neurons in all regions of prefrontal cortex (cingulate, agranular/insular and orbital cortices) and all cortical regions adjacent to prefrontal cortex, such as frontal, parietal and piriform cortex. Substantial number of D4 receptor-positive neurons has also been observed within the striatum and nucleus accumbens. In general, a clear stratification of the D4 receptor-positive neurons was observed in the cortex with the highest density seen in layers II/III and V/VI. D4 immunopositive material was also found in the dendritic processes, particularly clearly visible in the layer II/III. At the cellular level D4 receptor immunoreactivity was seen predominantly on the periphery of the cell body, but a certain population of neurons with clear cytoplasmatic localization was also identified. In addition to cortical distribution of D4 receptor-positive neurons we tried also to define types of neurons expressing D4 receptor protein. In double-labeling experiments, D4 receptor protein was found in nonphosphorylated neurofilament H-positive, calbindin-D28k-positive, as well as parvalbumin-positive cells. Since, used proteins are markers of certain populations of pyramidal neurons and GABA-ergic interneurons, respectively, our data indicate that D4 receptors are located on cortical pyramidal output neurons and their dendritic processes as well as on interneurons. Above localization indicates that D4 receptors are not only directly influencing excitability of cortical inter- and output neurons but also might be engaged in dendritic spatial and temporal integration, required for the generation of axonal messages. Additionally, our data show that D4 receptors are widely distributed throughout the cortex of rat brain, and that their cortical localization exceeds the localization of dopaminergic terminals.

DOI, an agonist of 5-HT2A/2C serotonin receptor, alters the expression of cyclooxygenase-2 in the rat parietal cortex.

The hallucinogenic effect of DOI, serotonin 5-HT2A/2C receptor agonist, is known to be associated with the activation of cortical 5-HT2 receptors. However, the effect of DOI on excitability of cortical neurons and their subsequent function is still not quite understood. Previous immunohistochemical studies using Fos proteins expression as a marker of neuronal activity showed the involvement of arachidonic acid cascade, particularly cyclooxygenase metabolic pathway, in DOI-induced Fos proteins expression in the rat parietal cortex. DOI increases arachidonic acid release which is transformed itself via acceleration of cyclooxygenase metabolic pathway to biologically active metabolites, such as prostaglandins and thromboxanes. Since cyclooxygenase-2 (COX-2) expression correlates with neuronal activity, it was of interest to investigate whether DOI is capable of influencing the level of COX-2 protein and mRNA expression in the rat parietal cortex. It was observed that neurons which were positive for 5-HT2A receptors showed constitutive COX-2 immunoreactivity. It was found further, that COX-2 protein level was increased at 1 h, and returned to the control level at 3 and 6 h after DOI (5 mg/kg) administration. In contrast, DOI decreased the COX-2 mRNA expression at all tested time points (1 h, 3h and 6h after DOI treatment). The obtained results further support the suggestion that COX-2 activation and possibly arachidonic acid metabolites generated by COX-2 may be considered as important mediators of functional responses generated by activation of cortical 5-HT2A/2C receptors.

Protection of dogs against canine distemper by vaccination with a canarypox virus recombinant expressing canine distemper virus fusion and hemagglutinin glycoproteins.

OBJECTIVES: To evaluate the safety and efficacy of a live canarypox virus recombinant-canine distemper virus (CDV) combination vaccine against virulent CDV challenge exposure, and to document lack of interference among the other modified-live virus (MLV) components. ANIMALS: 33 specific-pathogen-free (SPF) Beagle pups (7 to 10 weeks old). PROCEDURE: A canarypox virus recombinant-CDV combination vaccine was tested for safety and efficacy along with MLV components (canine adenovirus type 2, canine coronavirus, canine parainfluenza virus, and canine parvovirus) in 26 SPF Beagle pups. The combination vaccine was rehydrated with either Leptospira canicola-L icterohaemorrhagiae combination bacterin (vaccine 1) or sterile diluent (vaccine 2). An additional group of 7 seronegative SPF pups received the control MLV components devoid of the combination vaccine (vaccine 3). Two vaccinations were administered 21 days apart, either IM or SC. The dose of the combination vaccine used to inoculate these pups was 40 times lower than the recommended commercial dose. At 21 days after the booster vaccination, all pups were challenge exposed with a virulent CDV strain, then were observed for 21 days to record morbidity and mortality. RESULTS: Adverse local or generalized reactions were not induced by vaccinations. All vaccinates seroconverted to CDV. Serum antibody titers to MLV components were not different, with or without inclusion of the combination vaccine. After challenge exposure, morbidity and mortality in vaccinates were 0% (0/26); in control dogs, values were 100% morbidity and 86% mortality (6/7). Brain impression smear slides made from all dogs that did not survive challenge exposure were CDV positive by use of a direct fluorescein isothiocyanate method. CONCLUSIONS: The canarypox virus-CDV combination vaccine, administered SC or IM, is a safe product that elicits CDV seroconversion, does not interfere with other vaccine components, and protects vaccinated pups against virulent CDV challenge exposure.

Structure-activity relationship studies of CNS agents. Part 10(1): 1-Aryl-2-[3-(4-aryl-1-piperazinyl)propyl]-1,4-dihydro-3(2H)-isoquino linones: two modes of the interaction with the 5-HT1A receptor site.

The synthesis and 5-HT1A and 5-HT2 receptor affinities of 1-aryl-2-[3-(4-aryl-1-piperazinyl)propyl]-1,4-dihydro-3(2H)- isoquinolinones 7-28 are reported. The two derivatives 7 and 13 were the most potent 5-HT1A ligands (Ki 1.72 +/- 0.07 and 2.75 +/- 0.59 nM, respectively) of all the investigated compounds. It has been found that the effect of the substituent in the 1-arylpiperazine portion is opposite to the observed in simple 1-arylpiperazine. The molecular modelling results indicate that the investigated derivatives may interact with 5-HT1A sites in two different ways: as ordinary 4-substituted 1-arylpiperazines, or in such a manner that the aryl substituent at position 1 of the 3(2H)-isoquinolinone moiety and N-4 piperazine atom mimics remarkably well the bioactive conformation of simple 1-arylpiperazines.