Immunoreactive Fibers Research Articles

Leptin Receptor Expression in GABAergic Cells is Not Sufficient to Normalize Metabolism and Reproduction in Mice.

Previous studies indicate that leptin receptor (LepR) expression in GABAergic neurons is necessary for the biological effects of leptin. However, it is not clear whether LepR expression only in GABAergic neurons is sufficient to prevent the metabolic and neuroendocrine imbalances caused by LepR deficiency. In the present study, we produced mice that express the LepR exclusively in GABAergic cells (LepRVGAT mice) and compared them with wild-type (LepR+/+) and LepR-deficient (LepRNull/Null) mice. Although LepRVGAT mice showed a pronounced reduction in body weight and fat mass, as compared with LepRNull/Null mice, male and female LepRVGAT mice exhibited an obese phenotype relative to LepR+/+ mice. Food intake was normalized in LepRVGAT mice; however, LepRVGAT mice still exhibited lower energy expenditure in both sexes and reduced ambulatory activity in the females, compared with LepR+/+ mice. The acute anorexigenic effect of leptin and hedonic feeding were normalized in LepRVGAT mice despite the hyperleptinemia they present. Although LepRVGAT mice showed improved glucose homeostasis compared with LepRNull/Null mice, both male and female LepRVGAT mice exhibited insulin resistance. In contrast, LepR expression only in GABAergic cells was sufficient to normalize the density of agouti-related peptide (AgRP) and α-MSH immunoreactive fibers in the paraventricular nucleus of the hypothalamus. However, LepRVGAT mice exhibited reproductive dysfunctions, including subfertility in males and alterations in the estrous cycle of females. Taken together, our findings indicate that LepR expression in GABAergic cells, although critical to the physiology of leptin, is insufficient to normalize several metabolic aspects and the reproductive function in mice.

Another controller system for arterial pressure. AngII-vasopressin neural network of the parvocellular paraventricular nucleus may regulate arterial pressure during hypotension

Angiotensin II (AngII) immunoreactive cells, fibers and receptors, were found in the parvocelluar region of paraventricular nucleus (PVNp) and AngII receptors are present on vasopressinergic neurons. However, the mechanism by which vasopressin (AVP) and AngII may interact to regulate arterial pressure is not known. Thus, we tested the cardiovascular effects of blockade of the AngII receptors on AVP neurons and blockade of vasopressin V1a receptors on AngII neurons. We also explored whether the PVNp vasopressin plays a regulatory role during hypotension in anesthetized rat or not. Hypovolemic-hypotension was induced by gradual bleeding from femoral venous catheter. Either AngII or AVP injected into the PVNp produced pressor and tachycardia responses. The responses to AngII were blocked by V1a receptor antagonist. The responses to AVP were partially attenuated by AT1 antagonist and greatly attenuated by AT2 antagonist. Hemorrhage augmented the pressor response to AVP, indicating that during hemorrhage, sensitivity of PVNp to vasopressin was increased. By hemorrhagic-hypotension and bilateral blockade of V1a receptors of the PVNp, we found that vasopressinergic neurons of the PVNp regulate arterial pressure towards normal during hypotension. Taken together these findings and our previous findings about angII (Khanmoradi and Nasimi, 2017a) for the first time, we found that a mutual cooperative system of angiotensinergic and vasopressinergic neurons in the PVNp is a major regulatory controller of the cardiovascular system during hypotension.

The Role of Neuropeptide Y in the Nucleus Accumbens.

Neuropeptide Y (NPY), an abundant peptide in the central nervous system, is expressed in neurons of various regions throughout the brain. The physiological and behavioral effects of NPY are mainly mediated through Y1, Y2, and Y5 receptor subtypes, which are expressed in regions regulating food intake, fear and anxiety, learning and memory, depression, and posttraumatic stress. In particular, the nucleus accumbens (NAc) has one of the highest NPY concentrations in the brain. In this review, we summarize the role of NPY in the NAc. NPY is expressed principally in medium-sized aspiny neurons, and numerous NPY immunoreactive fibers are observed in the NAc. Alterations in NPY expression under certain conditions through intra-NAc injections of NPY or receptor agonists/antagonists revealed NPY to be involved in the characteristic functions of the NAc, such as alcohol intake and drug addiction. In addition, control of mesolimbic dopaminergic release via NPY receptors may take part in these functions. NPY in the NAc also participates in fat intake and emotional behavior. Accumbal NPY neurons and fibers may exert physiological and pathophysiological actions partly through neuroendocrine mechanisms and the autonomic nervous system.

Exogenous orexin-A downregulates luteinizing hormone secretory activity in prepubertal female rats.

Orexin-A is a neuropeptide synthesized in the lateral hypothalamus. Orexin-A immunoreactive fibres overlap distribution with GnRH neurons. In adult rats, orexin A is known to affect LH secretion via GnRH release modulation. Because data concerning the impact of orexin-A on the hypothalamo-pituitary axis activity are limited, we focused on the involvement of orexin-A and receptors of NPY in the modulation of LH release and LH subunit b (Lhb) mRNA expression in prepubertal female rats. Forty immature female Wistar rats were divided into 4 groups and received 2 intracerebroventricular (icv) microinjections of: 1 - artificial cerebrospinal fluid (CSF) (controls); 2 - CSF followed by orexin A; 3 - selective NPY receptor antagonist (BIBP) followed by CSF; 4 - BIBP followed by orexin A. One hour after the last microinjection, all rats were decapitated. Trunk blood was collected, and serum was stored at -20°C for the LH RIA examination. The adenohypophysis was immediately excised, flash-frozen, and kept at -80°C for RNA extraction. Real-time PCR amplification was carried out, and relative Lhb gene expression was calculated. In comparison to the CSF-treated controls with a mean LH serum concentration of 0.40 ± 0.02 ng/mL, the mean LH serum level was diminished both after orexin-A (0.27 ± 0.01 ng/mL) and after BIBP (0.30 ± 0.02 ng/mL) icv microinjections. In the presence of BIBP, orexin-A more effectively inhibited LH release (0.20 ± 0.01 ng/mL) when compared to the BIBP-treated group. Orexin-A and BIBP exerted a consistent inhibitory effect on Lhb mRNA expression levels in the anterior pituitary gland. In comparison to the CSF-treated controls, orexin-A, and BIBP-treated females responded with, respectively, 35% and 40% reduction of Lhb mRNA expression. Orexin-A and BIBP co-administration evoked a further reduction of Lhb gene transcriptional activity. Orexin-A exerts a down-regulatory effect on LH synthesis and release in immature female rats. Considering that Y1R-oriented down-regulation of endogenous NPY activity did not reverse the suppressive effect of exogenous orexin-A, it might be suggested that NPY and orexin A systems can operate independently to affect gonadotropin activity in the anterior pituitary of the immature female rats.

Morphology of the digestive system in carnivorous freshwater dourado Salminus brasiliensis.

The digestive system of teleost shows remarkable functional and morphological diversity. In this study, the digestive tract and accessory organs of dourado Salminus brasiliensis are characterized using anatomical, histological, histochemical and immunohistochemical analyses. The existence of taste buds bordered by microridges in the oesophagus of dourado was recorded for the first time, thus showing that the species drives food intake by either swallowing or rejecting the food item. The Y-shaped stomach of dourado consisted of cardiac, cecal and pyloric regions with tubular gastric glands registered solely in the cardiac and cecal segments. The intestine is a short N-shaped tube with two loops, an intestinal coefficient of 0.73. The structure of pyloric caeca is similar to that of the intestine wall, comprising tunica mucosa, tela submucosa, tunica muscularis and tunica serosa layers. Histochemical analyses revealed an increased incidence of goblet cells from the midgut to the hindgut segment. A well-developed enteric plexus of scattered nerve cell and fibres are found along the digestive tract, and the calcitonin gene-related peptide (CGRP) immunoreactive neurons and fibres were identified in the myenteric plexus from the oesophagus to the hindgut. The exocrine pancreas appears diffuse in the mesentery around the stomach, intestine and also reaches the liver, and the endocrine pancreas is organized as a few islets of Langerhans. The liver comprises three distinct, asymmetric lobes, and the portal triad arrangement was registered in this tissue.

Melatonin mediates monochromatic light-induced expression of somatostatin in the hypothalamus and pituitary of chicks

Melatonin (MEL) plays an important role in regulating growth and development of organisms and the cellular metabolism. This study was conducted to explore the role of MEL in mediating monochromatic light-induced secretion of somatostatin (SST) in the hypothalamus and pituitary in chicks. Pinealectomy models of newly hatched broilers were exposed to white (WL), red (RL), green (GL), and blue (BL) lights. The results showed that SST immunoreactive neurons and fibers were distributed in the hypothalamus. SST and SST receptor 2 (SSTR2) mRNA and protein levels in the hypothalamus and pituitary were higher in chicks exposed to RL than in chicks exposed to GL and BL. However, after pinealectomy, the mRNA and protein levels of SST and SSTR2 in the hypothalamus and pituitary in the different light groups were increased, and the differences between the groups disapeared. The expression trend of SSTR5 mRNA in the pituitary was the idential to that of SSTR2 mRNA in the pituitary. In vitro, exogenous SST inhibited growth hormone (GH) secretion, and selective antogonists of SSTR2 and SSTR5 promoted GH secretion. Selective antogonists of the melatonin receptor 1b (Mel1b) and Mel1c increased the relative concentrations of SST in the adenohypophysis cells. These results indicated that monochromatic light affects the expression of SST in chick hypothalamus and pituitary. MEL, via Mel1b and Mel1c, decreased SST secretion under GL, which was associated with the inhibition of SST, SSTR2, and SSTR5 in adenohypophysis cells.

Testosterone Treatment Mimics Seasonal Downregulation of Dopamine Innervation in the Auditory System of Female Midshipman Fish

In seasonally breeding vertebrates, hormones coordinate changes in nervous system structure and function to facilitate reproductive readiness and success. Steroid hormones often exert their effects indirectly via regulation of neuromodulators, which in turn can coordinate the modulation of sensory input with appropriate motor output. Female plainfin midshipman fish (Porichthys notatus) undergo increased peripheral auditory sensitivity in time for the summer breeding season, improving their ability to detect mates, which is regulated by steroid hormones. Reproductive females also show differences in catecholaminergic innervation of auditory circuitry compared with winter, non-reproductive females as measured by tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholaminergic synthesis. Importantly, catecholaminergic input to the inner ear from a dopaminergic-specific forebrain nucleus is decreased in the summer and dopamine inhibits the sensitivity of the inner ear, suggesting that gonadal steroids may alter auditory sensitivity by regulating dopamine innervation. In this study, we gonadectomized non-reproductive females, implanted them with estradiol (E2) or testosterone (T), and measured TH immunoreactive (TH-ir) fibers in auditory nuclei where catecholaminergic innervation was previously shown to be seasonally plastic. We found that treatment with T, but not E2, reduced TH-ir innervation in the auditory hindbrain. T-treatment also reduced TH-ir fibers in the forebrain dopaminergic cell group that projects to the inner ear, and likely to the auditory hindbrain. Higher T plasma in the treatment group was correlated with reduced-ir TH terminals in the inner ear. These T-treatment induced changes in TH-ir fibers mimic the seasonal downregulation of dopamine in the midshipman inner ear and provide evidence that steroid hormone regulation of peripheral auditory sensitivity is mediated, in part, by dopamine.

Daily and Estral Regulation of RFRP-3 Neurons in the Female Mice.

Female reproductive success relies on proper integration of circadian- and ovarian- signals to the hypothalamic-pituitary-gonadal axis in order to synchronize the preovulatory LH surge at the end of the ovarian follicular stage with the onset of the main active period. In this study, we used a combination of neuroanatomical and electrophysiological approaches to assess whether the hypothalamic neurons expressing Arg-Phe amide-related peptide (RFRP-3), a gonadotropin inhibitory peptide, exhibit daily and estrous stage dependent variations in female mice. Furthermore, we investigated whether arginine vasopressin (AVP), a circadian peptide produced by the suprachiamatic nucleus regulates RFRP-3 neurons. The number of c-Fos–positive RFRP-3 immunoreactive neurons is significantly reduced at the day-to-night transition with no difference between diestrus and proestrus. Contrastingly, RFRP neuron firing rate is higher in proestrus as compared to diestrus, independently of the time of the day. AVP immunoreactive fibers contact RFRP neurons with the highest density observed during the late afternoon of diestrus and proestrus. Application of AVP increases RFRP neurons firing in the afternoon (ZT6-10) of diestrus, but not at the same time point of proestrus, indicating that AVP signaling on RFRP neurons may depend on circulating ovarian steroids. Together, these studies show that RFRP neurons integrate both daily and estrogenic signals, which downstream may help to properly time the preovulatory LH surge.

Hericium erinaceus mycelium and its small bioactive compounds promote oligodendrocyte maturation with an increase in myelin basic protein

Oligodendrocytes (OLs), myelin-producing glia in the central nervous system (CNS), produce a myelin extension that enwraps axons to facilitate action potential propagation. An effective approach to induce oligodendrogenesis and myelination is important to foster CNS development and promote myelin repair in neurological diseases. Hericium (H.) erinaceus, an edible and culinary-medicinal mushroom, has been characterized as having neuroprotective activities. However, its effect on OL differentiation has not yet been uncovered. In this study using oligodendrocyte precursor cell (OPC) cultures and an ex vivo cerebellar slice system, we found that the extract from H. erinaceus mycelium (HEM) not only promoted the differentiation of OPCs to OLs in the differentiation medium, but also increased the level of myelin basic protein (MBP) on neuronal fibers. Moreover, daily oral administration of HEM into neonatal rat pups for 7 days enhanced MBP expression and OLs in the corpus callosum of the postnatal rat brain. The effect of HEM-derived bioactive compounds, the diterpenoid xylosides erinacine A (HeA) and HeC and a sesterterpene with 5 isoprene units called HeS, were further evaluated. The results showed that HeA and HeS more potently stimulated MBP expression in OLs and increased the number of OLs. Moreover, overlap between MBP immunoreactivity and neuronal fibers in cultured cerebellar tissue slices was significantly increased in the presence of HeA and HeS. In summary, our findings indicate that HEM extract and its ingredients HeA and HeS display promising functional effects and promote OL maturation, providing insights into their potential for myelination in neurodevelopmental disorders.

Identification and localization of a gonadotropin-releasing hormone-related neuropeptide in Biomphalaria, an intermediate host for schistosomiasis.

Freshwater snails of the genus Biomphalaria serve as obligatory hosts for the digenetic trematode Schistosoma mansoni, the causative agent for the most widespread form of intestinal schistosomiasis. Within Biomphalaria, S. mansoni larvae multiply and transform into the cercariae form that can infect humans. Trematode development and proliferation is thought to be facilitated by modifications of host behavior and physiological processes, including a reduction of reproduction known as "parasitic castration." As neuropeptides participate in the control of reproduction across phylogeny, a neural transcriptomics approach was undertaken to identify peptides that could regulate Biomphalaria reproductive physiology. The present study identified a transcript in Biomphalaria alexandrina that encodes a peptide belonging to the gonadotropin-releasing hormone (GnRH) superfamily. The precursor and the predicted mature peptide, pQIHFTPDWGNN-NH2 (designated Biom-GnRH), share features with peptides identified in other molluscan species, including panpulmonates, opisthobranchs, and cephalopods. An antibody generated against Biom-GnRH labeled neurons in the cerebral, pedal, and visceral ganglia of Biomphalaria glabrata. GnRH-like immunoreactive fiber systems projected to all central ganglia. In the periphery, immunoreactive material was detected in the ovotestis, oviduct, albumen gland, and nidamental gland. As these structures serve crucial roles in the production, transport, nourishment, and encapsulation of eggs, disruption of the GnRH system of Biomphalaria could contribute to reduced reproductive activity in infected snails.

Parasympathetic neurons in the human submandibular ganglion

The submandibular ganglion (SMG) contains parasympathetic neurons which innervate the submandibular gland. In this study, immunohistochemistry for vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), choline acetyltransferase (ChAT), dopamine β-hydroxylase (DBH), tyrosine hydroxylase (TH), and the transient receptor potential cation channel subfamily V members 1 (TRPV1) and 2 (TRPV2) was performed on the human SMG. In the SMG, 17.5 % and 8.9 % of parasympathetic neurons were immunoreactive for VIP and TRPV2, respectively. SMG neurons mostly contained ChAT- and DBH-immunoreactivity. In addition, subpopulations of SMG neurons were surrounded by VIP (69.6 %)-, TRPV2 (54.4 %)- and DBH (9.5 %)-immunoreactive (-ir) nerve fibers. SMG neurons with pericellular VIP- and TRPV2-ir nerve fibers were significantly larger than VIP- and TRPV2-ir SMG neurons, respectively. Other neurochemical substances were rare in the SMG. In the human submandibular gland, TRPV1- and TRPV2-ir nerve fiber profiles were seen around blood vessels. Double fluorescence method also demonstrated that TRPV2-ir nerve fiber profiles were located around myoepithelial and acinar cells in the submandibular gland. VIP and TRPV2 are probably expressed by both pre- and post-ganglionic neurons innervating the submandibular and sublingual glands. VIP, DBH and TRPV2 may have functions about regulation of salivary components in the salivary glands and neuronal activity in the SMG.

Visualizing the Calcitonin Gene-Related Peptide Immunoreactive Innervation of the Rat Cranial Dura Mater with Immunofluorescence and Neural Tracing.

The aim of this study was to examine the distribution and origin of the calcitonin gene-related peptide (CGRP)-immunoreactive sensory nerve fibers of the cranial dura mater using immunofluorescence, three-dimensional (3D) reconstruction and retrograde tracing technique. Here, the nerve fibers and blood vessels were stained using immunofluorescence and histochemistry techniques with CGRP and fluorescent phalloidin, respectively. The spatial correlation of dural CGRP-immuoreactive nerve fibers and blood vessels were demonstrated by 3D reconstruction. Meanwhile, the origin of the CGRP-immunoreactive nerve fibers were detected by neural tracing technique with fluorogold (FG) from the area around middle meningeal artery (MMA) in the cranial dura mater to the trigeminal ganglion (TG) and cervical (C) dorsal root ganglia (DRGs). In addition, the chemical characteristics of FG-labeled neurons in the TG and DRGs were also examined together with CGRP using double immunofluorescences. Taking advantage of the transparent whole-mount sample and 3D reconstruction, it was shown that CGRP-immunoreactive nerve fibers and phalloidin-labeled arterioles run together or separately forming a dural neurovascular network in a 3D view, while the FG-labeled neurons were found in the ophthalmic, maxillary, and mandibular branches of TG, as well as the C2-3 DRGs ipsilateral to the side of tracer application in which some of FG-labeled neurons presented with CGRP-immunoreactive expression. With these approaches, we demonstrated the distributional characteristics of CGRP-immunoreactive nerve fibers around the blood vessels in the cranial dura mater, as well as the origin of these nerve fibers from TG and DRGs. From the perspective of methodology, it may provide a valuable reference for understanding the complicated neurovascular structure of the cranial dura mater under the physiological or pathological condition.

Structural basis for noradrenergic regulation of neural circuits in the mouse olfactory bulb.

Olfactory input is processed in the glomerulus of the main olfactory bulb (OB) and relayed to higher centers in the brain by projection neurons. Conversely, centrifugal inputs from other brain regions project to the OB. We have previously analyzed centrifugal inputs into the OB from several brain regions using single-neuron labeling. In this study, we analyzed the centrifugal noradrenergic (NA) fibers derived from the locus coeruleus (LC), because their projection pathways and synaptic connections in the OB have not been clarified in detail. We analyzed the NA centrifugal projections by single-neuron labeling and immunoelectron microscopy. Individual NA neurons labeled by viral infection were three-dimensionally traced using Neurolucida software to visualize the projection pathway from the LC to the OB. Also, centrifugal NA fibers were visualized using an antibody for noradrenaline transporter (NET). NET immunoreactive (-ir) fibers contained many varicosities and synaptic vesicles. Furthermore, electron tomography demonstrated that NET-ir fibers formed asymmetrical synapses of varied morphology. Although these synapses were present at varicosities, the density of synapses was relatively low throughout the OB. The maximal density of synapses was found in the external plexiform layer; about 17% of all observed varicosities contained synapses. These results strongly suggest that NA-containing fibers in the OB release NA from both varicosities and synapses to influence the activities of OB neurons. The present study provides a morphological basis for olfactory modulation by centrifugal NA fibers derived from the LC.

Virus Injection to the Pituitary via Transsphenoidal Approach and the Innervation of Anterior and Posterior Pituitary of Rat.

The theory holds that the anterior pituitary in mammals receives humoral regulation. Previous studies have reported that the pars distalis of the anterior pituitary of several mammalian species contains substance P-, calcitonin gene-related peptide (CGRP)-, and galanin-like immunoreactive nerve fibers, but the origins of these nerve fibers are unclear. Removal of the pituitary gland, also called hypophysectomy, involves methods that access the pituitary gland via the transauricular or parapharyngeal pathways. However, these methods are not applicable for viral tracer injection to investigate the innervation of the anterior pituitary. The transauricular technique leads to inaccuracies in locating the pituitary gland, while the parapharyngeal approach causes high mortality in animals. Here, we introduce a protocol that accesses the pituitary gland in the rat via the transsphenoidal pathway. This method imitates surgical manipulations such as endotracheal intubation and sphenoid bone drilling, which involve the use of custom-made devices. Using the transsphenoidal pathway greatly improves the survival rate of rats because no additional dissection of blood vessels and nerves is required. Moreover, the pituitary gland can be viewed clearly and directly during the operation, making it possible to accurately inject pseudorabies virus (PRV) 152-expressing enhanced green fluorescent protein (EGFP) into the anterior or posterior pituitary, respectively. After injecting PRV 152 into the anterior pituitary, we found no evidence of direct innervation of the anterior pituitary in the rat brain. However, PRV 152 injection into the posterior pituitary revealed retrograde transneuronal cell bodies in many brain areas, including the CA1 field of the hippocampus, the basolateral amygdaloid nucleus, posterior part (BLP), the arcuate hypothalamic nucleus (Arc), the dorsal portion of the dorsomedial hypothalamic nucleus (DMD), the suprachiasmatic nucleus (SCh), and the subfornical organ (SFO). In the present study, we provide a description of a possible model of hypophysectomy or pituitary injection, and identify brain regions involved in regulating the rat pituitary gland using transneuronal retrograde cell body labeling with PRV.

Morphological elucidation of short-chain fatty acid receptor GPR41-positive enteric sensory neurons in the colon of mice with dextran sulfate sodium-induced colitis

Although the etiology of inflammatory bowel disease (IBD) remains unclear, it has generally been accepted that abnormalities in the intestinal immune system and dysbiosis of the gut microbiota are involved in the pathology of IBD. Recently, short-chain fatty acids (SCFAs) produced by gut microbiota were reported to maintain intestinal homeostasis through their receptors, such as GPR41. However, there are contradictory reports about the role of GPR41 in intestinal inflammation. Consequently, the roles of GPR41 in dysbiosis induced by intestinal inflammation remain unclear. Thus, we investigated the distribution of GPR41 in the colonic mucosa of mice with dextran sulfate sodium (DSS)-induced colitis. GPR41-immunoreactive fibrous structures were observed in the colonic lamina propria and muscularis layer of normal mice. In addition, GPR41-immunoreactive fibrous structures partly colocalized with calcitonin gene-related peptide (CGRP; a neurotransmitter of cholinergic enteric sensory neurons)-immunoreactive nerve fibers in the colonic lamina propria, indicating that GPR41 is expressed in cholinergic intrinsic sensory neurons. Furthermore, both GPR41-immunoreactivities and CGRP-immunoreactivities were significantly increased in the lamina propria of the colon in mice with DSS-induced colitis. Interestingly, GPR41-immunoreactivities were often found in close proximity to F4/80+ macrophages in the colonic mucosa of normal mice, and their frequency was elevated in the colonic mucosa of mice with DSS-induced colitis. Therefore, the crosstalk between SCFA-sensing intrinsic sensory neurons and macrophages might be involved in the pathology of acute colitis.

Co-transplantation of Schwann cells and neural stem cells in the laminin-chitosan-PLGA nerve conduit to repair the injured recurrent laryngeal nerve in SD rats.

The objective of this paper is to investigate the possibility and efficacy of recurrent laryngeal nerve repair by transplantation of co-cultured Schwann cells and neural stem cells (NSCs) in laminin-chitosan-poly-lactic-co-glycolic acid (laminin-chitosan-PLGA) nerve conduits in rats. A laminin-chitosan-PLGA conduit was used in a rat recurrent laryngeal nerve transection model. The rat recurrent laryngeal nerve was dissected to generate a 5 mm defect. Then, a laminin-chitosan-PLGA nerve conduit with or without Schwann cells and NSCs in the lumen was transplanted into the defect. A total of 96 female rats were randomised into six groups: co-culture of NSCs and Schwann cells in the nerve conduit group (CO), Schwann cells only in the nerve conduit group (SC), neural stem cells only in the nerve conduit group (NSC-only), nerve conduit group (null), autologous nerve graft group (autograft) and sham operation group (sham). Regenerated nerves were evaluated by histological and functional assessment at 8 and 12 weeks after surgery. The diameter and area of the regenerated myelin sheath, as well as the secretion of brain-derived neurotrophic factor and glial cell-derived neurotrophic factor in laryngeal muscle or regenerated nerve tissue in the CO group, were significantly better than they were in the SC, NSC-only and null groups (all P values < 0.05). Immunofluorescence showed that the CO group had significantly more neurofilament-200 immunoreactive and S-100 immunoreactive fibres than the SC, NSC-only and null groups (all P values < 0.05). The performance of the CO groups and autograft groups was found to be similar by laryngoscopy. Arytenoid cartilage motion recovery in these two groups was significantly better than it was in the other groups (all P values < 0.05). Our results indicated that co-culture of Schwann cells and NSCs in laminin-chitosan-PLGA conduits might promote injured nerve regeneration. This method might be a promising alternative for defective nerve repair.

The Chemokine CXCL14-like Immunoreactivity Co-exists with Somatostatin, but not NPY in the Rat Dorsal Horn and Has Intimate Association with GABAergic Neurons in the Lateral Spinal Nucleus.

Recent studies have proposed that the chemokine CXCL14 not only has a chemotactic activity, but also functions as a neuromodulator and/or neurotransmitter. In this study, we investigated the distribution of CXCL14 immunoreactive structures in the rat spinal cord and clarified the association of these structures with somatostatin, glutamic acid decarboxylase (GAD; a marker for GABAergic neurons), and neuropeptide Y (NPY). CXCL14 immunoreactive fibers and puncta were observed in lamina II, which modulates somatosensation including nociception, and the lateral spinal nucleus of the spinal dorsal horn at cervical, thoracic, and lumber spinal cord levels. These CXCL14 immunoreactive structures were also immuno-positive for somatostatin, but were immuno-negative for GAD and NPY. In the cervical lateral spinal nucleus, CXCL14 immunoreactive puncta, which were also immuno-positive for somatostatin, existed along the proximal dendrites of some of GABAergic neurons. Together, these results suggest that CXCL14 contributes to the modulation of somatosensation in concert with somatostatin. Neurons targeted by the CXCL14 fiber system include GABAergic neurons located in the lateral spinal nucleus suggesting that CXCL14 with somatostatin can influence the GABAergic neuron function.

Role of Substance P in the Pathophysiology of Inflammatory Bowel Disease and Its Correlation With the Degree of Inflammation.

Inflammatory bowel disease (IBD) is a multi-factorial, chronic inflammation of the gastrointestinal tract, containing ulcerative colitis (UC) and Crohn's disease (CD). In UC, inflammation and sores are confined morphologically and microscopically to the mucosa, the innermost surface of the colon and the rectum. Although, in CD, the infection is granulomatous and transmural, affecting the entire gastrointestinal tract from the mouth to the anus, with the skip area in-between. A Neuropeptide, substance P (SP), which acts as a neurotransmitter and as a neuromodulator, plays a vital role in the brain-gut axis under stress. Owing to the pro-inflammatory effects of SP, neuropeptide dysregulation induces inflammation in the intestine. There are variations in the distribution of substance P immunoreactive fibres in the various intestinal layers. The highest concentration of SP is in the mucosa and the lowest concentration in the lamina propria of the intestinal muscular membrane. Reduced vasoactive intestinal peptide (VIP) levels and elevated SP levels observed in the colonic mucosa of IBD by using immunohistochemistry and immunoassay. This literature review aims to find out the correlations between the level of substance P (SP) and disease activity. We conducted a literature review on IBD, SP, and we searched PubMed and Google Scholar for relevant articles in English. The result of the study supports a positive relationship between the level of substance P (SP) and disease activity, with increased concentration of substance p in the colon and rectum of CD and UC patients. It is concluded that patients with active CD, along with inflammatory changes, had elevated plasma SP levels and immunoreactivity of SP in the colon than those seen in control and inactive cases. These alterations are more prevalent in ulcerative colitis than Crohn's disease and are more prevalent in the moderately infected area than the least affected area of the intestine.

Chemical-neuroanatomical organization of peripheral sensory-efferent systems in the pond snail (Lymnaea stagnalis)

Perception and processing of chemical cues are crucial for aquatic gastropods, for proper elaboration of adaptive behavior. The pond snail, Lymnaea stagnalis, is a model species of invertebrate neurobiology, in which peripheral sensory neurons with different morphology and transmitter content have partly been described, but we have little knowledge regarding their functional morphological organization, including their possible peripheral intercellular connections and networks. Therefore the aim of our study was to characterize the sensory system of the tentacles and the lip, as primary sensory regions, and the anterior foot of Lymnaea with special attention to the transmitter content of the sensory neurons, and their relationship to extrinsic elements of the central nervous system. Numerous bipolar sensory cells were demonstrated in the epithelial layer of the peripheral organs, displaying immunoreactivity to antibodies raised against tyrosine hydroxylase, histamine, glutamate and two molluscan type oligopeptides, FMRFamide and Mytilus inhibitory peptide. A subepithelial plexus was formed by extrinsic serotonin and FMRFamide immunoreactive fibers, whereas in deeper regions axon processess of different origin with various immunoreactivities formed networks, too. HPLC–MS assay confirmed the presence of the low molecular weight signal molecules in the three examined areas. Following double-labeling immunohistochemistry, close arrangements were observed, formed by sensory neurons and extrinsic serotonergic (and FMRFamidergic) fibers at axo-dendritic, axo-somatic and axo-axonic levels. Our results suggest the involvement of a much wider repertoire of signal molecules in peripheral sensory processes of Lymnaea, which can locally be modified by central input, hence influencing directly the responses to environmental cues.

Effects of Different n6/n3 PUFAs Dietary Ratio on Cardiac Diabetic Neuropathy.

We studied the influence of experimentally induced DM1, in combination with different dietary n6:n3 polyunsaturated fatty acid (PUFA) ratios on different types of nerve fibers in rat myocardium, in order to reveal whether protective/unfavorable effects of different PUFAs on myocardial function in diabetic patients could be a (partial) repercussion of their effect on the changes in cardiac innervation. The control group (c) and diabetic group (stz) were fed with an n6/n3 ratio of ≈7; the diet of the stz+n6 group had an n6/n3 ratio ≈60, while the diet for the stz+DHA group contained 2.5% of fish oil (containing 16% eicosapentaenoic acid—EPA and 19% docosahexaenoic acid—DHA), n6/n3 ratio of ≈1. DM1 was induced by i.p. injection of streptozotocin (55 mg/kg) and rats were euthanized 30 days after induction. Immunohistochemistry was used for the detection and quantification of different types of neuronal fibers in the cardiac septum. We found changes in cardiac innervations characteristics for the initial phase of experimental DM1, which manifested as an increase in total number and area density of all neuronal fibers, measured by Pgp9.5 immunoreactivity. By detailed analysis, we found that this increase consisted mostly of heavy myelinated NF200 immunoreactive fibers and TH immunoreactive sympathetic fibers, while the density of ChAT immunoreactive parasympathetic fibers decreased. In the deep (middle) part of the myocardium, where rare fibers (of all studied types) were found, significant differences were not found. Surprisingly, we found a more consistent protective effect of n6 PUFAs, in comparison to n3 PUFAs supplementation. These results may provide a better understanding of the potential impacts of different PUFA ratios in the diet of diabetic patients on cardiac innervation and genesis and outcome of diabetic autonomic cardiomyopathy.