Immunoreactivity of acetylcholinesterase and M1 muscarinic receptors in the hippocampus and striatum of rats treated with Rebaudioside A.

Rebaudioside A (RebA) is a steviol glycoside used for production of sweeteners. It was shown that the glycosides affect memory and learning processes. The aim of the study was to investigate neurons immunoreactive for acetylcholinesterase (AChE) and M1 muscarinic receptors (mAChRs-M1) of the hippocampal CA1 and CA3 fields and striatal caudateputamen (CP) and globus pallidus (GP) in rats receiving RebA. RebA was administrated to adult rats for 45 days in dilutions of 1 mg and 2 mg RebA/ml water. Indirect peroxidase-antiperoxidase immunohistochemical reaction was conducted on frontal sections containing the hippocampus and striatum with use of antibodies against AChE and mAChRs-M1. Immunoreactive for the studied proteins neurons were morphologically and morphometrically assessed in hippocampal CA1 and CA3 fields and in CP and GP. Microscopic observations did not reveal significant changes in morphology of immunoreactive neurons, which suggests no neurotoxic effect of the studied glycoside on these cells. Morphometric analyses revealed an increase in the density of AChE and mAChRs-M1 immunoreactive neurons. A decrease in reaction intensity of AChE-positive neurons was also demonstrated in the hippocampal CA1 field and in GP. In contrast, an increase in reaction intensity of mAChRs-M1-positive neurons was found in CA1, CA3 fields and in CP and GP. The results of our preliminary studies indicate that RebA administrated to rats has an impact on cholinergic neurons in the studied area. The results suggest a possible increase in the activity of the cholinergic system, responsible for memory and learning processes, after administration of RebA.

Colocalization pattern of cocaine- and amphetamine-regulated transcript peptide and parvalbumin immunoreactivity in the hippocampus proper of the chinchilla.

This study set out to investigate, for the first time, the distribution and colocalization pattern of cocaine-and amphetamine-regulated transcript (CART) and one of the calcium binding- -proteins: parvalbumin (PV) in the chinchilla's hippocampus proper (HP). HP, consisting of Ammon's horn (CA) and the dentate gyrus (DG), is an important component of the limbic system, involved in learning and memory processes. CA showed a higher immunoreactivity of CART (-IR) compared to DG. CART-IR neurons were mainly observed in the molecular layer of DG and in the pyramidal layer of CA. CART-IR fibers were present in the granular layer; in the hilus numerous mossy fibers were detected, while in the molecular layer CART-IR fibers were not found. In all CA fields (CA1-CA3), CART-IR fibers were only present in the lacuno- sum-molecular layer. Immunofluorescence with double- labeling showed that only CART-IR cells stained positive for PV, whereas in CART-IR fibers there was no PV-positive reaction. Our research supplements missing knowledge about the distribution and colocalization pattern of CART with PV in the chinchilla's hippocampus, and also provides a better understanding of the similarities and differences among individuals of the same species and also with other mammals.

Immunoreactivity for calretininin interneurons of the hippocampal CA1 field and dentate gyrus in adult rats after administration of habanero peppers (Capsicum chinense Jacq.).

Calretinin (CR), a calcium-binding protein from EF-hand family, is localised in non-pyramidal GABA-ergic interneurons of the hippocampus. CR takes part in maintaining calcium binding homeostasis, which suggests its neuroprotective role. Hippocampal neurons contain membrane transient receptor potential vanilloid 1 (TRPV1) which binds to capsaicin (CAP) contained in habanero pepper fruits. Few in vivo studies have revealed the effect of CAP on interneurons containing CR. The aim of the present study was to investigate the CR immunoreac- tivity in interneurons of the hippocampal CA1 field and dentate gyrus (DG) in adult rats after intragastric admin- istration of the habanero pepper fruits. Wistar rats received a peanut oil - control group (C), and oil suspension of habanero pepper fruits at doses of 0.025 g dm/kg b.w. - group I and 0.08 g dm/kg b.w. - group II for 28 days. After euthanasia, the brains were collected and embedded in paraffin blocks using a routine histological tech- nique. Frontal hippocampal sections were immunohistochemically stained for CR by using a peroxidase-antiper- oxidase method. CR immunoreactive (CR-IR) interneurons were morphologically and morphometrically ana- lyzed under a light microscope. The results showed similar shapes and distribution of cells in both areas of the brain in group C and I of animals. However, CR-IR interneurons in the hippocampal CA1 field and in DG were occasionally observed in the group II of rats. The results of morphometric studies did not reveal statistically significant differences in the surface area and shape index of cells between examined brain regions from groups I and II compared to group C. Only in group II of rats, an increase in the digital immunostaining intensity of CR-IR interneurons was found in DG. Low number of CR-IR interneurons in the hippocampal CA1 field and in the DG, under the influence of a large dose of habanero pepper fruits containing CAP, may be caused by the activation of TRPV1 receptors and the increase in Ca2+ ions in these cells. This phenomenon may ultimately lead to neuronal death and may disturb neuronal conduction.

Effects of maternal treatment with ß-hydroxy-ß-metylbutyrate and 2-oxoglutaric acid on femur development in offspring of minks of the standard dark brown type.

The aim of the study was to evaluate the effect of the diet, mother type and sex of the offspring on the mechanical and geometric parameters of long bones as well as bone tissue density in minks. Primiparous and multiparous dams were supplemented with ß-hydroxy ß-methylbutyrate (a metabolite of leucine, at the daily dosage of 0.02 g/kg of body weight) and/or 2-oxoglutaric acid (a precursor of glutamine, at the daily dosage of 0.4 g/kg of body weight) during gestation. The diet did not influence bone tissue density and the length of the humerus. An increase in the length of the femur was noted in male offspring delivered by multiparous dams. The diet resulted in an increase in the weight of the humerus in males from multiparous dams and a decrease in offspring from primiparous dams. Heavier femora were noted in male offspring delivered by both types of dams. The maximum elastic strength of the humerus was higher in the offspring delivered by multiparous than primiparous dams, irrespective of the offspring sex. The diet resulted in reduction in the ultimate strength of the femur in the male offspring delivered by primiparous dams. Only females born by multiparous dams, irrespective of the diet, showed a significant increase in the cross-sectional area of the humerus, while a significant decline was noted in males delivered by multiparous dams and in all the offspring delivered by primiparous dams. An increase in the cross-sectional area of the femur was noted in the offspring delivered by multiparous dams, while reduction was observed in the offspring delivered by primiparous dams. These results have shown for the first time that the presence of ß-hydroxy-ß-methylbutyrate or 2-oxoglutaric acid in the diet of pregnant primiparous or multiparous dams unambiguously affects the geometry and mechanical properties of offspring's long bones.

Immunoreactivity of arcuate nucleus astrocytes in rats after intragastric administration of habanero peppers (Capsicum Chinese Jacq.).

Habanero pepper fruits contain capsaicin (CAP) characterised by a spicy taste. Astrocytes express vanilloid receptor (TRPV1), which interacts with cannabinoids including CAP. Only a few studies revealed that CAP leads to alterations of the arcuate nucleus (ARC) structures. The aim of this study was to analyse the GFAP (GFAP-IR) and S100ß (S100ß-IR) immunoreactive astrocytes of ARC in adult rats after intragastric administration of habanero pepper fruits. Adult, Wistar rats received a peanut oil - control group (C) - and oil suspension of habanero pepper fruits at a dose of 0.08 g dm/kg b.w. for 7 days - E1 group - and 28 days - E2 group. After euthanasia, the brains were embedded in paraffin blocks using a routine histological technique. Frontal slices of ARC were immunohistochemically stained for GFAP and S100ß using specific antibodies in the peroxidase-antiperoxidase method. Astrocytes of ARC were morphologically and morphometrically analysed under a light microscope. The results of the study did not reveal statistically significant changes in the density of GFAP-IR cells in E1 and E2 groups of rats in comparison with group C. A statistically significant increase in the density of S100ß-IR astrocytes was observed in the E1 group and a decrease in the E2 group. Astrocytes with expression of both studied proteins were characterised by morphological alterations in ARC in the E2 group. The obtained results suggest an influence of CAP contained in habanero pepper fruits on the reactivity of astroglia, which may have an impact on the astrocyte-neuron interactions in order to maintain a proper activity of nervous cells in ARC.

Immunohistochemical evaluation of hippocampal CA1 region astrocytes in 10-day-old rats after monosodium glutamate treatment.

High concentration of glutamate (Glu) is excitotoxic for nervous system structures. This may lead to glial reactivity ie. increased expression of glial fibrillary acidic protein (GFAP) and S100ß protein, and also to hypertrophy and proliferation of cells which are determined by the presence of Ki-67 antigen. The aim of the study was to analyse the immunoreactivity of the GFAP, S100ß and Ki-67 proteins in astrocytes of hippocampal CA1 region in young rats after administration of monosodium glutamate (MSG) at two doses: 2 g/kg b.w. (I group) and 4 g/kg b.w. (II group). In rats from I and II group morphologically altered astrocytes with the GFAP expression were observed in the SLM of the hippocampal CA1 region. The cells had eccentrically located nuclei and on the opposite site of the nuclei there were single or double, long and weakly branched processes. Moreover, in the SLM the increase of the number of GFAP and S100ß immunopositive astrocytes and nuclei with Ki-67 expression, in contrary to control individuals, was observed. These results suggest the increased expression of the proteins in early reactions or hyperplasia which, together with cell hypertrophy, indicate late reactivity of astroglia in response to glutamate noxious effect.