Behavior and severity of adjuvant arthritis in four rat strains.

Previous research has suggested that behavioral traits of the histocompatible Lewis and Fischer strains of rats could be related to the difference in their susceptibility to adjuvant arthritis (AA). In the present study, the predictive value of behavioral markers in susceptibility to AA was investigated in nonhistocompatible inbred DA, Lewis, Albino Oxford (AO), and outbred Wistar strain. Behavioral profiles (open filed test and forced swim test) were determined prior to immunization with a single intradermal injection of complete Freund's adjuvant. Animals were daily scored for clinical signs of AA. The occurrence of certain behaviors and clinical indices of AA was significantly associated with strain membership. Discriminant analysis identified strain-related behavioral and illness profiles with very few overlaps among the phenotypes. Discriminant classification significantly exceeded the proportion of cases, which could have been correctly classified on the basis of chance. Open field behavior, in particular, exploration and grooming, differentiated among AA-susceptible and AA-resistant strains. Multiple regression analysis indicated that severity of AA (maximum clinical sign) can be predicted by the latency time and grooming behavior in the open field independently of strain membership. No clear distinction between AA-susceptible and AA-resistant strains was found with respect to forced swim test immobility. It was concluded that (a) strain-related genetic predisposition is important for the expression of certain behavioral traits and for susceptibility to AA and (b) open field behaviors, particularly grooming and latency, predict susceptibility to AA across different rat strains.

Production of the Fos protein after contextual fear conditioning of C57BL/6N mice.

Male C57BL/6N mice were chosen to determine Fos production during acquisition of context-dependent fear and after re-exposure to the conditioning context. Fear-conditioning was induced by a single exposure of mice to a context followed by an electric shock. Control groups consisted of mice exposed to context only (Context group) or to an immediate electric shock. When contextual retention was measured 24 h after conditioning (retention test 1), significant contextual generalization was observed. However, when animals were exposed to a different context from days 2-5 after conditioning and then tested for retention on day 6 (retention test 2), generalization was markedly reduced. After the training, the fear-conditioned mice produced higher Fos levels than mice exposed to an immediate shock in the hippocampus, medial amygdaloid nucleus and parietal somatosensory cortex. Both shock groups produced significantly more Fos than the Context group in the central nucleus of the amygdala. After retention test 1, fear-conditioned mice generated more Fos in the hippocampus and central amygdaloid nucleus than the two control groups. However, all groups exhibited similarly low Fos production after retention test 2. The results demonstrated that simultaneous Fos production in the hippocampus, central and medial nuclei of amygdala and somatosensory parietal cortex closely paralleled the ability of mice to acquire conditioned fear. In contrast, Fos production after the retention tests did not correlate with the expression of conditioned fear.

Immunologic tolerance to myelin basic protein decreases stroke size after transient focal cerebral ischemia.

Immune mechanisms contribute to cerebral ischemic injury. Therapeutic immunosuppressive options are limited due to systemic side effects. We attempted to achieve immunosuppression in the brain through oral tolerance to myelin basic protein (MBP). Lewis rats were fed low-dose bovine MBP or ovalbumin (1 mg, five times) before 3 h of middle cerebral artery occlusion (MCAO). A third group of animals was sensitized to MBP but did not survive the post-stroke period. Infarct size at 24 and 96 h after ischemia was significantly less in tolerized animals. Tolerance to MBP was confirmed in vivo by a decrease in delayed-type hypersensitivity to MBP. Systemic immune responses, characterized in vitro by spleen cell proliferation to Con A, lipopolysaccharide, and MBP, again confirmed antigen-specific immunologic tolerance. Immunohistochemistry revealed transforming growth factor beta1 production by T cells in the brains of tolerized but not control animals. Systemic transforming growth factor beta1 levels were equivalent in both groups. Corticosterone levels 24 h after surgery were elevated in all sham-operated animals and ischemic control animals but not in ischemic tolerized animals. These results demonstrate that antigen-specific modulation of the immune response decreases infarct size after focal cerebral ischemia and that sensitization to the same antigen may actually worsen outcome.

Changes in immunological and neuronal conditions markedly altered antibody response to intracerebroventricularly injected ovalbumin in the rat.

The serum antibody response to ovalbumin (OA) has been investigated following intracerebroventricular (i.c.v.) and intravenous administration of antigen in the rat, under altered neuronal and immunological conditions. I.c.v. administration of antigen was far more potent in eliciting humoral immune response. Central nervous system (CNS) immunization under the conditions of disrupted blood-brain barrier decreased anti-OA antibody production. Peripheral polyclonal stimulation with Bordetella pertussis increased production of specific antibodies to i.c.v. injected antigen, while complete Freund's adjuvant had no effect on the immune response. These results suggest that CNS compartmentalization of antigen may be critical for mounting strong antibody production, and that peripheral polyclonal stimulation of the immune system may markedly contribute to the overall intensity of the immune response.

Stress-induced rise in serum anti-brain autoantibody levels in the rat.

Sera from Wistar rats subjected to different stress procedures were tested by ELISA for the presence of autoantibodies with specificity for neuron-specific enolase (NSE) and S100 protein that are preferentially localized in neurons and glia, respectively. Autoantibodies were present in sera of animals before exposure to stress, and raised with age. Anti-NSE and anti-S100 autoantibody levels were increased one day after termination of restraint (2 hours daily, 10 days) and electric tail shock (80 shocks daily, 19 days), and in fifth and tenth week of overcrowding stress. Differences between stressed and control animals were not present one month following restraint and electric tail shock and in twentieth week of overcrowding.

Experimental allergic encephalomyelitis in adult DA rats subjected to neonatal handling or gentling.

The present study investigated the effect of daily handling and gentling between postnatal days 1 and 28 on experimental allergic encephalomyelitis (EAE) in 8-week old DA rats. Handling consisted of removing pups from the mother, and placing them in the novel cage for 15 min. The gentling procedure included handling accompanied by 3 min of dorsal tactile stimulation before returning the pups to the nest cage. Adult rats of both sexes handled in infancy showed increased susceptibility to EAE, as revealed by higher incidence of the disease, and more severe clinical signs. Anti-myelin basic protein (MBP) autoantibodies were increased in handled males, and decreased in handled females, compared to controls. Gentling induced aggravation of clinical signs and histopathological lesions of EAE in males, while in gentled females suppression was observed. These results indicated that both neonatal handling and gentling aggravated EAE induced in adult male rats. In female rats handling exacerbated, and gentling suppressed clinical EAE. The overall effect of neonatal manipulations was more pronounced in males. Furthermore, in mothers separated from their offspring due to handling and gentling, and immunized for EAE at day 28 postpartum, earlier appearance of clinical signs, and increased frequency of relapses compared to control dams was recorded.

Suppression of experimental allergic encephalomyelitis in offspring of DA and Wistar rats following immunization of mother with encephalitogen.

Immunization of female rats with encephalitogen before gestation, during gestation, and during lactation differentially decreased susceptibility to experimental allergic encephalomyelitis (EAE) in their offspring. The most pronounced suppression, revealed by lowered incidence and weaker clinical signs of the disease, was observed in offspring of mothers immunized before gestation and during lactation in both Dark August (EAE-susceptible), and Wistar (EAE-relatively resistant) rat strains. Induction of EAE in mothers during pregnancy only delayed the onset of the disease in DA progeny. The overall effect on EAE in offspring did not depend on the disease intensity in mothers. Our results suggest that anti-myelin basic protein (MBP) antibodies passively transferred from mothers are not responsible for the observed protection in offspring.

Maternal deprivation and early weaning modulate experimental allergic encephalomyelitis in the rat.

The present experiment deals with the effect of maternal deprivation (MD) and early weaning (EW) on the development and course of experimental allergic encephalomyelitis (EAE) in Dark August (DA) rats. Five litters (five to nine pups per liter) were subjected to MD (4 h daily) from Day 1 until Day 28. EW rats were weaned on Day 15 (EW-15, five litters) or Day 21 (EW-21, four litters). Control rats and MD rats were weaned on Day 28. At the age of 8 weeks, rats were immunized with guinea pig spinal cord in complete Freund's adjuvant and clinical signs of EAE were recorded daily. On Day 18 after immunization, rats were bled and sacrificed. Brain and spinal cord were examined histologically for EAE lesions. Serum anti-rat myelin basic protein (MBP) antibodies were detected by ELISA. MD female rats exhibited suppression of neurological and histological signs of EAE in comparison with control rats. MD and control females showed elevated anti-MBP antibody level compared to MD and control males. EW-15 female rats demonstrated potentiation of neurological signs of EAE compared to control females. EW-21 females developed more severe clinical signs and histological lesions compared to control females. These results show that neonatal experiences, such as maternal deprivation and early weaning, influence the development of EAE in adult DA rats.

The site of action of Ca2+ in the activation of steroidogenesis: studies in Ca(2+)-clamped bovine adrenal zona-glomerulosa cells.

The Ca(2+)-messenger system plays a crucial role in the regulation of steroid production in adrenal zona-glomerulosa cells, as it is known to mediate the action of both angiotensin II and K+. In the present study we used intact isolated glomerulosa cells in which the cytosolic free Ca2+ concentration ([Ca2+]c) was clamped at various levels with the Ca2+ ionophore ionomycin in order to locate the site(s) of action of Ca2+. By measuring in parallel steroid synthesis and [Ca2+]c, we show that Ca2+ levels (50-860 nM) regulate the production of both pregnenolone (up to 669 +/- 71.1% of the basal production) and aldosterone (up to 301 +/- 42.2%; EC50 = 303 nM). By contrast, Ca2+ did not stimulate the conversion of 11-deoxycorticosterone into aldosterone. Ca2+ modulation did not affect the formation of pregnenolone from freely diffusible analogues of cholesterol, indicating that Ca2+ acts at a step upstream of cholesterol side-chain cleavage. Moreover cycloheximide, an inhibitor of protein translation and of adrenocorticotropin-induced facilitation of intramitochondrial cholesterol transport, the rate-limiting step in steroidogenesis, also blocked Ca(2+)-triggered pregnenolone formation. This is consistent with a model in which Ca2+ promotes cholesterol transfer between mitochondrial membranes. In addition, agents using the cyclic AMP pathway as well as angiotensin II potentiated the steroidogenic response to increases in [Ca2+]c by augmenting both the efficacy and the potency of Ca2+. This effect of angiotensin II did not involve protein kinase C. These results establish a direct link between agonist-induced [Ca2+]c rises and a specific step of the steroidogenic pathway.

Effect of Met-enkephalin and opioid antagonists on rat macrophages.

Effects of Met-enkephalin (Met-ENK) and opioid antagonists on H2O2 release by peritoneal macrophages from DA and AO rats were investigated. Met-ENK increased and decreased H2O2 production by macrophages of DA and AO rats, respectively. These effects were antagonized by low, but not high, concentrations of naloxone and ICI 174864. High concentrations of both antagonists directly modulated H2O2 release and retained the strain-related differences seen with Met-ENK. The results showed direct, strain- and dose-dependent, effects of Met-ENK, naloxone, and ICI 174864 on rat macrophage function.

Neonatal sound stress and development of experimental allergic encephalomyelitis in Lewis and DA rats.

This experiment deals with the effect of neonatal sound stress on the susceptibility of rats in adult life to the induction of experimental allergic encephalomyelitis (EAE). Two inbred strains of rats, Lewis and DA, highly susceptible to EAE were used. On postnatal days 15, 18 and 21, animals of both sexes were sound stressed in a sound attenuated chamber (90dB, 60 rings/5 sec during 1 h, on a variable interval schedule) in the presence or absence of the mother. Experimental groups were as follows: (a) pups stressed without the mother (SP); (b) pups stressed in the presence of the mother (SPM); (c) control nonstressed pups separated from the mother (CP), and (d) control nonstressed pups undisturbed in their nest cages (CPM). Rats were weaned on postnatal day 28. At the age of 8 weeks, all groups were immunized with guinea pig spinal cord in complete Freund's adjuvant. Signs of EAE were recorded daily until the day 20 after immunization when animals were bled and sacrificed. Serial sections of cerebrum, cerebellum and spinal cord were examined histologically for the presence of mononuclear cell infiltrates. Anti-myelin basic protein (MBP) antibodies were detected in serum samples using ELISA technique. Stressed Lewis rats (groups SP and SPM) compared to control groups CP and CPM, developed more severe EAE as revealed by a higher aggregate clinical score, more pronounced histological lesions and increased production of anti-MBP antibodies. The presence of the mother during stress session (group SPM) prolonged the disease.(ABSTRACT TRUNCATED AT 250 WORDS)