Effects of in ovo injection of prebiotics and synbiotics on the productive performance and microstructural features of the superficial pectoral muscle in broiler chickens.

The aim of the study was to compare the effects of 2 prebiotics and 2 synbiotics injected in ovo on productivity parameters, quality, and microstructure of the superficial pectoral muscle in 35-day-old broiler chickens. On day 12 of incubation, 9,000 eggs Ross 308 were randomly divided into 5 experimental groups treated with different bioactives in ovo injected: C, control with physiological saline; PI, with 1.760 mg inulin; PB, with 0.528 mg of commercial prebiotic Bi2tos; SI, with 1.760 mg inulin and 1,000 CFU Lactococcus lactis spp. lactis IBB SL1; SB, with 0.528 mg Bi2tos and 1,000 CFU Lactococcus lactis spp. cremoris IBB SC1. The synbiotic solution contained 20 µl bacterial suspension and 180 µl prebiotic solution. For productive parameters and further tests ten male birds for each experimental group were used. The birds were slaughtered on day 35 of age. At slaughter, samples of the left pectoral muscles were taken and preserved by freezing in liquid nitrogen. The pH and color of the meat were evaluated at 45 min and 24 h post-mortem. Water holding capacity (WHC) was measured and expressed as the percentage of free water in meat. Microscopic specimens were analysed using MultiScan software for the measurement of the percentage of oxidative and glycolytic fibres and mean diameter of the muscle fibres. In ovo injection of prebiotics Bi2tos had a positive effect on body weight. In prebiotic group (PI) a negative impact on hatchability was observed. Prebiotics and synbiotics had no influence on the yield of the carcass and pectoral muscle. Bioactive compounds had a significant effect on the quality of meat parameters such as: pH 24 h (PI and PB group), L* 45' (SI and SB group), and WHC (groups PB, SI, and SB). The analysis of the enzymatic profile showed a significant increase in the percentage of glycolytic fibres in the pectoral muscle from chicken treated with a synbiotic with the addition of inulin (group SI).

Chronic stress changes prepulse inhibition after amphetamine challenge: the role of the dopaminergic system.

The goal of this research was to examine the influence of chronic mild stress (CMS) on prepulse inhibition (PPI). We used an amphetamine challenge to study the role of the dopaminergic system in limbic structures. Chronic stress caused a reduction in both sucrose preference and body weight. It was found that the initially strong response to amphetamine in the control rats was weakened after stress on both the behavioural and biochemical levels: improved PPI, decreased dopamine D2 receptor expression in the central nucleus of amygdala (CeA) and nucleus accumbens (NAC), and decreased dopamine and 3-MT (3-methoxytyramine) levels in NAC. We observed that the stress-evoked attenuation of amphetamine-induced stimulation was also paralleled by changes in corticosterone level. These effects were accompanied by a decrease in both glutamate and the glutamate/gamma-aminobutric acid (GABA) ratio in the NAC. The interpretation of these results is that prolonged stress induces compensatory mechanisms in the mesolimbic system which are responsible for psychostimulant (amphetamine) effects.

The impact of synbiotic administration through in ovo technology on the microstructure of a broiler chicken small intestine tissue on the 1st and 42nd day of rearing.

BACKGROUND: Application the innovative method which is in ovo technology provides a means of modulating the immune system at early embryonic stages. The aim of study was to determine influence of the in ovo stimulation, on d 12 of incubation, with synbiotics (synbiotic 1- L. salivarius IBB3154 + Bi2tos, Clasado Ltd. and the synbiotic 2 - L. plantarum IBB3036 + lupin RFOs) on the microstructure of duodenum, jejunum and ileum in the 1st and 42nd day of rearing. RESULTS: On the 1st day of chickens life, in the duodenum of both experimental groups (SYN1 and SYN2), a significantly higher and wider intestinal villi as well as a significantly larger absorbent surface of these villi were found in comparison with the Control group (P ≤ 0.01). On the 42nd day of rearing the beneficial effect of synbiotic 1 was reflected by the numerically higher villi (no statistical differences) with a larger surface (P ≤ 0.01) in the duodenum in the SYN1 group compare to the Control group. In the jejunum on the 1st day of life, in the SYN1 group, significantly higher villi than in the Control group, with a simultaneous decrease in the depth of crypts (P ≤ 0.01), and also the largest width of villi and their absorbent area (P ≤ 0.01) in comparison to the other groups were found. On the 42nd day of life, in the jejunum, an increase in the height of the villi whilst reducing the crypt depth in the SYN2 group was found (P ≤ 0.01). In turn, in the SYN1 group, there were significantly more neutral goblet cells observed compared with the control group (P ≤ 0.05). In the ileum of 1-day-old chickens, the widest villi (P ≤ 0.05) and the deepest crypts (P ≤ 0.01) were found in the SYN2 group. In the same group, there was also the least amount of neutral goblet cells in comparison to the other groups (P ≤ 0.05). CONCLUSIONS: We observed that synbiotic 1 and 2 beneficially affected the examined characteristics on the 1st and 42nd day of life. The obtained results allow us to conclude that the use of synbiotics significantly affect gut structure which should contribute to improvement in nutrient absorption by the gut.

Corticotropin releasing factor receptor 1 antagonist differentially inhibits freezing behavior and changes gamma-aminobutyric acidergic activity in the amygdala in low- and high-anxiety rats.

The aim of this study was to examine the effects of non-peptide corticotropin-releasing factor receptor 1 (CRF1) antagonist (antalarmin) administration on rat conditioned fear responses and gamma-aminobutyric acid (GABA)-ergic brain activity (GAD67 expression and GABA concentration) in low-anxiety (LR) and high-anxiety (HR) rats. The animals were divided into the LR and HR groups based on the duration of their conditioned freezing response in the first contextual fear test. After 28 days, the animals were re-subjected to the contextual fear training and test. The rats received an antalarmin injection (10 mg/kg or 20 mg/kg) 80 min before the second exposure to the aversive context. Antalarmin significantly attenuated the conditioned fear response only in the HR rats. The behavioral effect of a lower dose (10 mg/kg) of antalarmin was accompanied by increased GAD67 expression in the prelimbic cortex (PL) and central nucleus of the amygdala (CeA) and an increased GABA concentration in the amygdala. These studies showed that HR rats were more susceptible to the anxiolytic effects of CRF1 antagonist administration, which were associated with increased GABAergic activity in the medial prefrontal cortex and amygdala. The current data may provide insights into the neurobiological mechanism operating within the mesolimbic CRF-GABA neurotransmitter systems, which may be responsible for individual differences in stress-related diseases. This knowledge can be applied to further elucidate the pathophysiology of anxiety and trauma/stress-related disorders.

Is the interaction between fatty acids and tryptophan responsible for the efficacy of a ketogenic diet in epilepsy? The new hypothesis of action.

The effects of a ketogenic diet in controlling seizure activity have been proven in many studies, although its mechanism of action remains elusive in many regards. We hypothesize that the ketogenic diet may exert its antiepileptic effects by influencing tryptophan (TRP) metabolism. The aim of this study was to investigate the influence of octanoic and decanoic fatty acids (FAs), the main components in the MCT diet (medium-chain triglyceride diet, a subtype of the ketogenic diet), on the metabolism of TRP, the activity of the kynurenic pathway and the concentrations of monoamines and amino acids, including branched-chain amino acids (BCAA) and aromatic amino acids (AAA) in rats. The acute effects of FA on the sedation index and hippocampal electrical after-discharge threshold were also assessed. We observed that intragastric administration of FA increased the brain levels of TRP and the central and peripheral concentrations of kynurenic acid (KYNA), as well as caused significant changes in the brain and plasma concentrations of BCAA and AAA. We found that the administration of FA clearly increased the seizure threshold and induced sedation. Furthermore, we have demonstrated that blocking TRP passage into the brain abolished these effects of FA but had no similar effect on the formation of ketone bodies. Given that FAs are major components of a ketogenic diet, it is suggested that the anticonvulsant effects of a ketogenic diet may be at least partly dependent on changes in TRP metabolism. We also propose a more general hypothesis concerning the intracellular mechanism of the ketogenic diet.

The kynurenine pathway: a missing piece in the puzzle of valproate action?

The present study was designed to determine the role of the kynurenine pathway (KP) in the mechanism of action of valproate (VPA). Therefore, we investigated changes in the concentrations of tryptophan (TRP), kynurenic acid (KYNA), and kynurenine (KYN) in the brain and plasma following VPA administration (50, 250 and 500mg/kg i.p.). The most important findings of our study were that VPA administration produced a progressive and strong increase in the central concentrations of KYNA, KYN and TRP. Simultaneously, the TRP level in plasma declined, while the peripheral increase of KYNA in plasma was weaker and occurred earlier than in the hippocampus. Bearing in mind that the observed effect may be a result of a strong VPA-induced displacement of TRP from its binding sites to plasma albumin, we checked the effect of ibuprofen (IBU) administration (a prototypic drug used to study drug binding to serum albumin) on the KP. We found that IBU evoked a similar pattern of change in the KP activity as VPA. These new findings indicate the existence of a mechanism that could stimulate the production of KYNA in the brain after VPA administration, and may partially contribute to the mechanisms of VPA action. The results of our experiment indicate that an increase in the brain's KYNA level may be achieved by TRP displacement from its binding site on plasma albumin with the administration of different drugs, including VPA, IBU, or short-chain fatty acids, with important clinical consequences.

Corticosterone modulates fear responses and the expression of glucocorticoid receptors in the brain of high-anxiety rats.

The aim of our experiments was to assess the effect of acutely administered corticosterone on the expression of glucocorticoid receptors (GRs) in the brain of rats with high (HR) and low (LR) levels of anxiety. The rats were divided into groups according to their conditioned fear-induced freezing responses and then were subjected to a second conditioned fear session one week after the initial fear conditioning. Immunocytochemical analysis revealed that the second exposure to contextual aversive stimuli resulted in higher levels of GRs expression in cingulate cortex area 1 (Cg1), the secondary motor cortex (M2) of the prefrontal cortex and the dentate gyrus of the hippocampus (DG) in LR rats compared with HR rats. The pretreatment of HR rats with corticosterone (20mg/kg, sc) increased the expression levels of GRs in Cg1, the M2 area and the DG to the levels observed in the LR vehicle group. The increase in the GRs levels was accompanied by a significant decrease in the conditioned fear response in the HR group. The control animals that were not exposed to aversive stimuli had similar levels of receptor-related immunoreactivity in all brain regions, and corticosterone did not change these expression levels. Our results suggest that HR animals may have deficits in the expression of stress-induced GRs in the prefrontal cortex and the DG. In addition, pretreatment with corticosterone increases the expression of GRs and normalizes the fear response in HR rats.

Influence of in ovo prebiotic and synbiotic administration on meat quality of broiler chickens.

A trial was conducted to evaluate the effect of in ovo injection of prebiotic and synbiotics on growth performance, meat quality traits (cholesterol content, intramuscular collagen properties, fiber measurements), and the presence of histopathological changes in the pectoral muscle (PS) of broiler chickens. On d 12 of incubation, 480 eggs were randomly divided into 5 experimental groups treated with different bioactives, in ovo injected: C, control with physiological saline; T1 with 1.9 mg of raffinose family oligosaccharides; T2 and T3 with 1.9 mg of raffinose family oligosaccharides enriched with different probiotic bacteria, specifically 1,000 cfu of Lactococcus lactis ssp. lactis SL1 and Lactococcus lactis ssp. cremoris IBB SC1, respectively; T4 with commercially available synbiotic Duolac, containing 500 cfu of both Lactobacillus acidophilus and Streptococcus faecium with the addition of lactose (0.001 mg/embryo). Among the hatched chickens, 60 males were randomly chosen (12 birds for each group) and were grown to 42 d in collective cages (n = 3 birds in each 4 cages: replications for experimental groups). Broilers were fed ad libitum commercial diets according to age. In ovo prebiotic and synbiotic administration had a low effect on investigated traits, but depend on the kind of bioactives administered. Commercial synbiotic treatment (T4) reduced carcass yield percentage, and the feed conversion ratio was higher in T3 and T4 groups compared with other groups. The abdominal fat, the ultimate pH, and cholesterol of the PS were not affected by treatment. Broiler chickens of the treated groups with both slightly greater PS and fiber diameter had a significantly lower amount of collagen. The greater thickness of muscle fibers (not significant) and the lower fiber density (statistically significant), observed in treated birds in comparison with those of the C group, are not associated with histopathological changes in the PS of broilers. The incidence of histopathological changes in broiler chickens from examined groups was low, which did not affect the deterioration of meat quality obtained from these birds.

Corticosterone attenuates conditioned fear responses and potentiates the expression of GABA-A receptor alpha-2 subunits in the brain structures of rats selected for high anxiety.

The aim of the experiment was to assess the effects of an acutely administered corticosterone on the expression of GABA-A receptor alpha-2 subunits in the brain structures of high (HR) and low (LR) anxiety rats (divided according to their conditioned fear-induced freezing response) subjected to a second conditioned fear session (1 week after fear conditioning). We found that corticosterone (20 mg/kg, sc) given to rats prior to the second conditioned fear session significantly enhanced a decrease in fear expression in the HR group. The behavioural effect of fear was accompanied by the increased expression of alpha-2 subunits in the basolateral amygdala (BLA) and the dentate gyrus of the hippocampus (DG) of the HR group. Corticosterone potentiated the effect of fear on alpha-2 subunit expression in the BLA, DG, the cingulate cortex area 1 and the secondary motor cortex (areas Cg1 and M2). The current study provides insight into the mechanisms that may be responsible for the beneficial effects of glucocorticoids in the therapy of some anxiety disorders.

The effects of electrical hippocampal kindling of seizures on amino acids and kynurenic acid concentrations in brain structures.

Our study demonstrated that the development of seizures during the electrically induced kindling of seizures is associated with significant changes in the concentration of kynurenic acid (KYNA) and its precursor, tryptophan (TRP). The primary finding of our study was an increase in KYNA levels and the KYNA/TRP ratio (a theoretical index of activity of the kynurenine pathway) in the amygdala and hippocampus of kindled animals. We also found decreases in the concentration of tryptophan in the hippocampus and prefrontal cortex. Changes in the concentration of KYNA and TRP in the amygdala were accompanied by a significant decrease in γ-Aminobutryic Acid (GABA) levels and an increase in the glutamate/GABA ratio. Moreover, we found a significant negative correlation between the local concentrations of KYNA and glutamate in the amygdala of kindled rats. However, there were no changes in the local concentrations of the following amino acids: glutamate, aspartate, glutamine, glycine, taurine and alanine. In conclusion, these new results suggest a modulatory influence of KYNA on the process of epileptogenesis, characterized by a negative relationship between the KYNA and glutamate systems in the amygdala.

Expression of N-methyl-D-aspartate (R)(GluN2B) - subunits in the brain structures of rats selected for low and high anxiety.

In this paper, we studied differences in the density of N-methyl-D-aspartate (NMDA) receptor GluN2B subunits in the brains of low (LR) and high (HR) anxiety rats subjected to extinction trials and re-learning of a conditioned fear response, modeling a natural course of anxiety disorders. Classifications of animals as LR or HR was determined by fear-induced freezing responses in the contextual fear test. Increased basal concentrations of GluN2B subunits were observed in the amygdala of HR rats as compared to the unconditioned control group by Western blot analysis. Re-exposure of HR animals to the fear-conditioned context resulted in elevated concentrations of GluN2B subunits in the amygdala, hippocampus and the prefrontal cortex compared to LR rats as well as in the hippocampus and prefrontal cortex vs. the control group. In addition, it was shown that re-test of a conditioned fear increased the number of cells expressing GluN2B subunits in the basolateral amygdala, dentate gyrus of the hippocampus and secondary motor cortex (M2) in the HR group relative to the LR group. Together, these data suggest that animals that are more anxious have altered patterns of GluN2B subunit expression in the frontal cortex and limbic structures, which control emotional behaviour.

The effect of CRF2 receptor antagonists on rat conditioned fear responses and c-Fos and CRF expression in the brain limbic structures.

The influence of intracerebroventricular-administered selective corticotropin-releasing factor receptor 2 (CRF(2)) antagonists (antisauvagine-30, astressin-2B), on rat anxiety-like behavior, expression levels of c-Fos and CRF, and plasma corticosterone levels were examined in the present study. In fear-conditioned animals, both CRF receptor antagonists enhanced a conditioned freezing fear response and increased the conditioned fear-elevated concentration of serum corticosterone. Exogenously administered antisauvagine-30 increased the aversive context-induced expression of c-Fos in the 1 and 2 areas of the cingulate cortex (Cg1, Cg2), the central amygdala (CeA) and parvocellular neurons of the paraventricular hypothalamic nucleus (pPVN), and it enhanced the effect of conditioned fear in the secondary motor cortex (M2) and medial amygdala (MeA). Immunocytochemistry demonstrated an increase in CRF expression in the Cg1, M2 areas of the cortex, and pPVN, and it revealed the effect of conditioned fear in the CeA 35 min after antisauvagine-30 administration and 10 min after the conditioned fear test. Furthermore, astressin-2B, another CRF(2) receptor antagonist, enhanced expression of c-Fos and CRF in the CeA and pPVN, and revealed the effect of conditioned fear in the Cg1. These data support a model in which an excess in CRF(1) receptor activation, combined with reduced CRF(2) receptor signaling, may contribute to stronger expression of anxiety-like responses.

Gamma-aminobutyric acid concentrations in benign parotid tumours and unstimulated parotid saliva.

OBJECTIVE: Apart from its role as an inhibitory neurotransmitter, γ-aminobutyric acid is also thought to regulate various stages of cell proliferation and differentiation in the brain and periphery. The present study aimed to assess the levels of γ-aminobutyric acid and its biochemical precursor glutamic acid (glutamate) in benign parotid tumours and in unstimulated parotid saliva. METHOD: Unstimulated parotid saliva was collected bilaterally, using the swab method, in 20 patients with unilateral pleomorphic adenoma or Warthin's tumour. Samples of tumour and adjacent salivary tissue were collected during tumour resection. RESULTS: Concentrations of γ-aminobutyric acid and glutamate, but not aspartate, were significantly higher in the tumour tissue than in the non-tumour tissue. There was no significant difference in salivary concentrations of γ-aminobutyric acid, glutamate or aspartate, comparing the involved and non-involved side. CONCLUSION: The present results provide preliminary evidence that γ-aminobutyric acid may be involved in the growth of benign parotid tumours.

The localization of brain sites of anxiogenic-like effects of urocortin-2.

The influence of intracerebroventricullary-administered urocortin-2, a selective corticotropin-releasing factor receptor 2 (CRF(2)) agonist, on rat anxiety-like behaviour, the expression of c-Fos and CRF, and plasma corticosterone levels was examined in the present study. When applied to animals exposed to the conditioned fear-induced context, urocortin-2 enhanced a conditioned freezing fear response. Urocortin-2 also significantly decreased rat exploratory activity in the open field test. Exogenous urocortin-2 increased the conditioned fear-induced expression of c-Fos in the central amygdala (CeA), and parvocellular neurons of the paraventricular hypothalamic nucleus (pPVN), and revealed the effect of conditioned fear in the medial amygdala (MeA). In the fear-conditioned animals, immunocytochemistry showed an increase in the density of CRF-related immunoreactive complexes in the lateral septum (LS), 35min after urocortin-2 administration and 10min after the conditioned fear test, compared with saline-pretreated fear-conditioned animals. These data suggest a role of urocortin-2 in the behavioural and immunocytochemical responses to stress, in which it strengthens the measures of anxiety-like responses.

Role of autophagy in mammary gland development.

Autophagy is a highly conserved catabolic process responsible for degradation and recycling of long-lived proteins and organelles by lysosomes. This degradative pathway, together with proteasome system is particularly important during development and under certain environmental stress conditions. This review summarizes the latest achievements of studies aiming to explore the role of autophagy in development and differentiation of eukaryotic cells. It shows the importance of this process in the development of lower eukaryotic organisms such as Dicyostelium discoideum, and Caenorhabditis elegans, as well as functions of autophagy and autophagy related genes (Atg) in development and differentiation of higher eukaryotic organisms. The review is focused on the results of studies conducted on mammary gland, as it is a good model for studying the mechanisms controlling higher eukaryotic organisms' development. Studies have shown that autophagy is involved in the removal of epithelial cells during formation of alveolar structures, indicating its role in mammogenesis. There are also evidences of involvement of Atg's in epithelial tumors development. Context dependent manipulations of autophagic pathways may create more effective anticancer therapies in the future.

The opposite role of hippocampal mGluR1 in fear conditioning in kindled and non-kindled rats.

In the present paper, we analyzed the effects of hippocampal mGluR1 on the consolidation of a fear-conditioned response and on hippocampal glutamate and GABA concentration in rats subjected to the chemically-induced kindling of seizures. We hypothesized the important role of this glutamate receptor subpopulation in behavioural disturbances accompanying epilepsy. To this end, the behavioural and biochemical effects of selective mGluR1 and 5 receptor ligands were compared in sham and kindled animals (pentylenetetrazol-induced seizures). It was found that despite the fact that the freezing response to the aversively conditioned context was not changed by kindling itself, post-training intrahippocampal (dentate gyrus) injection of AIDA (a mGluR1 antagonist) oppositely influenced rat freezing behaviour in the non-kindled and kindled animals (i.e. the receptor ligand increased and decreased duration of the fear reaction, respectively). Kindling of seizures also enhanced the Glutamate/GABA ratio in the dorsal hippocampus (in vivo microdialysis), indicating an enhancement of excitatory processes in the brain. Altogether, the results showed that kindling of seizures led the potentiation of excitatory processes in the hippocampus, changing the role of the local mGluRs1 population in the conditioned fear learning.

[An analysis of the course of experimental pneumocystosis in rats using some serological methods]. / Analiza przebiegu doswiadczalnej pneumocystozy u szczurów za pomoca wybranych metod serologicznych.

Since serological methods used in the diagnosis of pneumocystosis may be helpful to a various extent, depending on the stage of a diagnosed infection, it was decided to evaluate the usefulness of some the methods in the course of infection. Wistar rats were used in the experiments. The animals were administered hydrocortisone injections for 12 weeks to induce immunosuppression and activate naturally occurring asymptomatic infections with Pneumocystis carinii. Blood samples and specimes of lung tissues were collected, then they were examined for the presence of specific IgG and IgM antibodies, circulating antigen of Pneumocystsis carinii and circulating immune complexes using immunoenzymatic assays. The results of the above experiments indicated, that in an early stage of infection, the examinations of serum samples for circulating immune complexes were helpful, particularly for these with IgM antibodies

[Nested PCR in the diagnosis of Pneumocystis carinii pneumonia (PCP) in AIDS patients]. / "Nested" PCR w diagnostyce przypadków pneumocystozowego zapalenia pluc (PCP) u pacjentów z AIDS.

The studies were undertaken to evaluate the usefulness of a nested PCR assay in the diagnosis of Pneumocystis carinii pneumonia (PCP) in AIDS patients. To achieve the end, 51 excretions samples from the respiratory tract were collected from HIV-infected patients with respiratory symptoms, and examined for the presence of specific DNA. A portion of the mitochondrial large-subunit rRNA gene of Pneumocystis carinii was amplified with outer primers pair pAZ 102E, pAZ 102H and internal primers pAZ 102X, pAZ 102Y. Positive nested PCR results were obtained with 36 out of 51 examined samples. Some 52.8% of the positive results were obtained with samples collected from patients without clinical diagnosis of PCP. It was concluded, that the nested PCR method, being too sensitive, is not suitable for the routine diagnosis of PCP in AIDS patients.