Leucin-enkefalin u pankreasu ovaca ima zajedničku izražajnost s tvari P, galaninom i somatostatinom

Leucine-enkephalin (Leu-Enk) is an endogenous opioid peptide that binds to opioid receptors. Leu-Enk is widely distributed in the central and peripheral nervous system. The aim of the present immunofluorescence study was to examine the distribution of Leu-Enk-immunoreactive (IR) neuronal elements in the ovine pancreas. Using double immunohistochemical staining, the co-localization of Leu-Enk with galanin, somatostatin and substance P was also studied. In the intrapancreatic ganglia, immunoreactivity to Leu-Enk was found in 64.9 ± 1.7% of neurons. Small arterioles and the ductal system were innervated by numerous Leu-Enk-IR nerve terminals. Moderate Leu-Enk-IR nerve fibres surrounded the islets of Langerhans but none of them penetrated into spaces between endocrine cells. In 66.7 ± 4.3% of Leu-Enk-immunoreactive intrapancreatic neurons, expression of galanin was found. A statistically smaller subpopulation of Leu-Enk-IR intrapancreatic neurons (37.4 ± 6.2%) exhibited immunoreactivity to SP. The expression of somatostatin was detected in the relatively smallest group (21.2 ± 3.8%) of Leu-Enk-positive intrapancreatic neurons. Co-expression of Leu-Enk and SP was detected in nerve terminals encircling the pancreatic small arterioles, connective tissue and ducts. Leu-Enk-positive nerve fibres around the islets of Langerhans were not immunoreactive for SP. None of the Leu-ENK-positive nerve fibres around the islets of Langerhans co-stored somatostatin. In general, there was also no co-localization between Leu-Enk and somatostatin in nerve terminals supplying small arterioles and veins. Co-expression of GAL and Leu-Enk was observed well in nerve fibres encircling the blood vessels, but not in nerve fibres of the connective tissue. We conclude that abundant immunoreactivity to Leu-Enk in the ovine pancreas and the co-localization of Leu-Enk with other regulatory neuropeptides may reflect the possible involvement of Leu-Enk as a regulator of the exocrine and endocrine pancreatic functions, as well as in regulation of pancreatic blood flow.Leucin-enkefalin (Leu-Enk) je endogeni opioidni peptid koji se veže na opioidne receptore. Leu-Enk je široko rasprostranjen u središnjem i perifernom živčanom sustavu. Cilj ovog istraživanja bio je pomoću imunofluorescencije utvrditi raspodjelu Leu-Enk-imunoreaktivnih (IR) neuronskih elemenata u pankreasu ovaca. Primjenom dvostrukog imunohistokemijskog bojenja, istražena je i zajednička lokalizacija Leu-Enk s galaninom, somatostatinom i tvari P. U intrapankreasnim ganglijima imunoreaktivnost prema Leu-Enku pronađena je u 64,9 ± 1,7% neurona. Male arteriole i kanalni sustav bili su inervirani mnogim Leu-Enk-IR živčanim završetcima. Umjerena Leu-Enk-IR živčana vlakna okruživala su Langerhansove otočiće, ali nijedno od njih nije prodrlo u prostore između endokrinih stanica. U 66,7 ± 4,3% Leu-Enk-imunoreaktivnih intrapankreasnih neurona otkrivena je ekspresija galanina. Statistički niža subpopulacija Leu-Enk-IR intrapankreasnih neurona (37,4 ± 6,2%) pokazala je imunoreaktivnost prema SP. Izražajnost somatostatina otkrivena je u relativno najmanjoj skupini (21,2 ± 3,8%) Leu-Enk-pozitivnih intrapankreasnih neurona. Zajednička izražajnost Leu-Enk i SP otkrivena je u živčanim završetcima koji okružuju male arteriole pankreasa, vezivno tkivo i kanale. Leu-Enk-pozitivna vlakna živaca oko Langerhansovih otočića nisu bila imunoreaktivna na SP. Nijedno od Leu-Enk-pozitivnih živčanih vlakana oko Langerhansovih otočića nije bilo zajednički lokalizirano sa somatostatin-imunoreaktivnim živčanim završetcima. Općenito, nije bilo zajedničke lokalizacije između Leu-Enka i somatostatin živčanih završetaka koji opskrbljuju male arteriole i vene. Zajednička izražajnost GAL i Leu-Enk dobro je vidljiva u živčanim vlaknima koja okružuju krvne žile, ali ne i u živčanim vlaknima vezivnog tkiva. Zaključeno je da obilna imunoreaktivnost Leu-Enk u pankreasu ovaca, te njegova zajednička lokalizacija s drugim regulatornim neuropeptidima, može odražavati moguću uključenost Leu-Enk u regulaciju egzokrine i endokrine funkcije pankreasa odnosno u regulaciju optjecaja krvi u pankreasu

Immunodetection of P2X2 Receptor in Enteric Nervous System Neurons of the Small Intestine of Pigs

Extracellular adenosine 5′-triphosphate (ATP) is one of the best-known and frequently studied neurotransmitters. Its broad spectrum of biological activity is conditioned by the activation of purinergic receptors, including the P2X2 receptor. The P2X2 receptor is present in the central and peripheral nervous system of many species, including laboratory animals, domestic animals, and primates. However, the distribution of the P2X2 receptor in the nervous system of the domestic pig, a species increasingly used as an experimental model, is as yet unknown. Therefore, this study aimed to determine the presence of the P2X2 receptor in the neurons of the enteric nervous system (ENS) of the pig small intestine (duodenum, jejunum, and ileum) by the immunofluorescence method. In addition, the chemical code of P2X2-immunoreactive (IR) ENS neurons of the porcine small intestine was analysed by determining the coexistence of selected neuropeptides, i.e., vasoactive intestinal polypeptide (VIP), substance P (sP), and galanin. P2X2-IR neurons were present in the myenteric plexus (MP), outer submucosal plexus (OSP), and inner submucosal plexus (ISP) of all sections of the small intestine (duodenum, jejunum, and ileum). From 44.78 ± 2.24% (duodenum) to 63.74 ± 2.67% (ileum) of MP neurons were P2X2-IR. The corresponding ranges in the OSP ranged from 44.84 ± 1.43% (in the duodenum) to 53.53 ± 1.21% (in the jejunum), and in the ISP, from 53.10 ± 0.97% (duodenum) to 60.57 ± 2.24% (ileum). Immunofluorescence staining revealed the presence of P2X2-IR/galanin-IR and P2X2-IR/VIP-IR neurons in the MP, OSP, and ISP of the sections of the small intestine. The presence of sP was not detected in the P2X2-IR neurons of any ganglia tested in the ENS. Our results indicate for the first time that the P2X2 receptor is present in the MP, ISP, and OSP neurons of all small intestinal segments in pigs, which may suggest that its activation influences the action of the small intestine. Moreover, there is a likely functional interaction between P2X2 receptors and galanin or VIP, but not sP, in the ENS of the porcine small intestine

Can Bioactive Compounds in Beetroot/Carrot Juice Have a Neuroprotective Effect? Morphological Studies of Neurons Immunoreactive to Calretinin of the Rat Hippocampus after Exposure to Cadmium

Cadmium ions (Cd2+) penetrate the blood–brain barrier and can, among other effects, influence intracellular calcium metabolism, leading to neurodegeneration. In the presented work, we estimated the effect of Cd2+ on the expression of calretinin in the neurons of the rat hippocampus and analyzed the reverse effect of freshly pressed beetroot/carrot juice in this context. In the 12-week lasting experiment, 32 8-week-old male Wistar rats were divided into four experimental groups (n = 8): the control group (C) received pure tap water; the Cd group (Cd)—received Cd2+ dissolved in tap water (5 mg Cd2+/kg b.w.); and two groups received beetroot/carrot juice: the BCJ group was administered only juice, and the Cd + BCJ group received juice with the addition of Cd2+ (5 mg Cd2+/kg b.w.). The exposition to low doses of Cd2+ caused a significant decrease in calretinin-immunoreactive (Cr-IR) neurons compared to the non-exposed groups. Moreover, the addition of Cd2+ to tap water reduced the numbers and length of Cr-IR nerve fibers. The negative effect of Cd2+ was significantly attenuated by the simultaneous supplementation of beetroot/carrot juice (Cd + BCJ). The study showed that the bioactive compounds in the beetroot/carrot juice can modulate Ca2+ levels in neurons, and thus, potentially act as a neuroprotective factor against neuronal damage

Magnoflorine from <i>Berberis vulgaris</i> Roots—Impact on Hippocampal Neurons in Mice after Short-Term Exposure

In search of novel potential drug candidates that could be used as treatments or prophylactics for memory impairment, an aporphine alkaloid magnoflorine (MAG) isolated from the root of Berberis vulgaris was proven to exhibit beneficial anti-amnestic properties. Its effects on immunoreactivity to parvalbumin in the mouse hippocampus were assessed together with a study on its safety and concentration in the brain and plasma. For this purpose, four experimental groups were created: the MAG10 group—treated with 10 mg MAG/kg b.w. i.p., the MAG20 group—treated with 20 mg MAG/kg b.w. i.p., the MAG50 group—treated with 50 mg MAG/kg b.w. i.p., and a control group—injected with saline i.p. at a volume corresponding to their weight. Our results indicated that the hippocampal fields CA1–CA3 were characterized by an elevated number of parvalbumin-immunoreactive neurons (PV-IR) and nerve fibers in mice at the doses of 10 and 20 mg/kg b.w. (i.p.). No significant changes to the levels of IL-1β, IL-6 or TNF-α were observed for the above two doses; however, the administration of 50 mg/kg b.w. i.p. caused a statistically significant elevation of IL-6, IL-1beta plasma levels and an insignificant raise in the TNF-alpha value. The HPLC–MS analysis showed that the alkaloid’s content in the brain structures in the group treated with 50 mg/kg b.w. did not increase proportionally with the administered dose. The obtained results show that MAG is able to influence the immunoreactivity to PV-IR in hippocampal neurons and might act as a neuroprotective compound

Basal Intestinal Morphology, Immunolocalization of Leptin and Ghrelin and Their Receptors in Newborn Wistar Rats after Prenatal Exposure to Fumonisins

Animal feed is very frequently contaminated with different types of mold, the metabolites of which are toxic to living organisms. Mold-contaminated cereal is rich in heat-resistant and harmful metabolites such as fumonisins (FBs). The amount of FBs consumed as part of animal feed, including livestock feed, is unknown. Therefore, this study aimed to evaluate the effects of maternal oral FB intoxication on basal duodenum morphology and the immunolocalization of gut hormones responsible for food intake (leptin and ghrelin), as well as their receptors, in newborn rat offspring. Pregnant Wistar rats were randomly allocated to one of three groups: a control group or one of two FB-intoxicated groups (60 or 90 mg FB/kg b.w., respectively). Basal morphological duodenal parameters changed in a dose- and sex-dependent manner. The intensity of the ghrelin immunoreaction was unchanged in females, while in males it increased after FB exposure (60 mg/kg b.w.), with a simultaneous decrease in expression of the ghrelin receptor. Leptin and its receptor immunoreaction intensity was decreased in both sexes following FB exposure. The current study highlighted the potential involvement of intestinal ghrelin and leptin in the metabolic disturbances observed later in life in offspring that were prenatally exposed to fumonisins

Morphology and Chemical Coding of Rat Duodenal Enteric Neurons following Prenatal Exposure to Fumonisins

Fumonisins (FBs), including fumonisin B1 and B2 produced by the fungus Fusarium verticillioides, are widespread mycotoxins contaminating crop plants as well as processed food. The aim of the experiment was to determine whether the exposure of 5-week-old pregnant rats to FBs at 60 mg/kg b.w. (group FB60) or 90 mg/kg b.w. (group FB90) results in morphological changes in the duodenum of weaned offspring, particularly the enteric nervous system (ENS). In addition, the levels of expression of galanin and vasoactive intestinal polypeptide (VIP) in the ENS were analysed by immunofluorescence in the control and experimental groups of animals. No significant morphological changes in the thickness of the muscle layer or submucosa of the duodenum were noted in group FB60 or FB90. In group FB90 (but not FB60), there was a significant increase in the width of the villi and in the density of the intestinal crypts. Immunofluorescence analysis using neuronal marker Hu C/D showed no significant changes in group FB60 or FB90 in the morphology of the duodenal ENS, i.e., the myenteric plexus (MP) and submucosal plexus (SP), in terms of the density of enteric ganglia in the MP and SP, surface area of MP and SP ganglia, length and width of MP and SP ganglia, surface area of myenteric and submucosal neurons, diameter of myenteric and submucosal neurons, density of myenteric and submucosal neurons, and number of myenteric and submucosal neurons per ganglion. In both groups, there was an increase (relative to the control) in the percentage of Hu C/D-IR/VIP-IR (IR-immunoreactive) and Hu C/D-IR/galanin-IR myenteric and submucosal neurons in the ganglia of both the MP and SP of the duodenum. In addition, in groups FB60 and FB90, there was an increase in the number of nerve fibres showing expression of VIP and galanin in the mucosa, submucosa and circular muscle layer of the duodenum. The results indicate that prenatal exposure to FBs does not significantly alter the histological structure of the duodenum (including the ENS) in the weaned offspring. The changes observed in the chemical code of the myenteric and submucosal neurons in both experimental groups suggest harmful activity of FBs, which may translate into activation of repair mechanisms via overexpression of neuroprotective neuropeptides (VIP and galanin)

Maternal acrylamide exposure changes intestinal epithelium, immunolocalization of leptin and ghrelin and their receptors, and gut barrier in weaned offspring

Abstract Acrylamide (ACR) is an amide formed as a byproduct in many heat-processed starchy-rich foods. In utero ACR exposure has been associated with restricted fetal growth, but its effects of postnatal functional development of small intestine is completely unknown. The current study investigated the time- and segment-dependent effects of prenatal ACR exposure on morphological and functional development of small intestine in weaned rat offspring. Four groups of pregnant female Wistar rats were exposed to ACR (3 mg/kg b.w./day) for 0, 5, 10 and 15 days during pregnancy. Basal intestinal morphology, immunolocalization of gut hormones responsible for food intake and proteins of intestinal barrier, activity of the intestinal brush border disaccharidases, apoptosis and proliferation in intestinal mucosa were analyzed in offspring at weaning (postnatal day 21). The results showed that in utero ACR exposure disturbs offspring gut structural and functional postnatal development in a time- and segment-depended manner and even a short prenatal exposure to ACR resulted in changes in intestinal morphology, immunolocalization of leptin and ghrelin and their receptors, barrier function, activity of gut enzymes and upregulation of apoptosis and proliferation. In conclusion, prenatal ACR exposure disturbed the proper postnatal development of small intestine

The Concentration of Selected Heavy Metals in Muscles, Liver and Kidneys of Pigs Fed Standard Diets and Diets Containing 60% of New Rye Varieties

The carry-over of heavy metals from feed to muscles is generally low if animals are fed with a standard diet containing amounts below the maximum permissible levels. However, prolonged exposure to heavy metals can lead to their accumulation in some organs like muscles, liver, and kidneys. This paves the way for human health risks related to the consumption of products of animal-origin. Thus, using feed mixtures with a low level of heavy metals in pig production will contribute to increasing public health and safety and is of environmental concern. The study aimed to assess the impact of the level of some heavy metals (Cd, Pb, Hg, Cu, Zn, Fe and Mn) in standard (control) feed mixtures and in alternative feed mixtures based on maize or new rye varieties (population and hybrid) on the heavy metal concentration in muscles, liver and kidney of fattened pigs at slaughter. While some differences between heavy metals content in examined tissue samples from experimental groups were observed, all of them were in the range of allowable levels according to European Community rules. In conclusion, new rye varieties, especially the hybrid variety, could be an alternative source of cereal grains for pig nutrition