Enteroendocrine profile of α-transducin and α-gustducin immunoreactive cells in the chicken (Gallus domesticus) gastrointestinal tract.

The enteroendocrine profile and distribution patterns of the taste signaling molecules, α-gustducin (Gαgust) and α-transducin (Gαtran) protein subunits, were studied in the gastrointestinal (GI) tract of the chicken (Gallus domesticus) using double labeling immunohistochemistry. Gαtran or Gαgust immunoreactivity was observed in enteroendocrine cells (EEC) expressing different peptides throughout the entire GI tract with different density. In the proventriculus tubular gland, Gαtran or Gαgust/gastrin (GAS) immunoreactive (-IR) cells were more abundant than Gαtran/or Gαgust containing glucagon-like peptide-1 (GLP-1) or peptide YY (PYY), whereas only few Gαtran or Gαgust cells co-stored ghrelin (GHR) or 5-hydroxytryptamine (5-HT). In the pyloric mucosa, many Gαtran or Gαgust-IR cells co-expressed GAS or GHR, with less Gαtran or Gαgust cells containing GLP-1, PYY, or 5-HT. In the small intestine, a considerable subset of Gαtran or Gαgust-IR cells co-expressed 5-HT in the villi of the duodenum and ileum, PYY in the villi of the jejunum, CCK or GLP-1 in the villi of the ileum, and GHR in the duodenum crypts. In the large intestine, many Gαtran or Gαgust-IR cells contained 5-HT or GLP-1 in the villi of the rectum, whereas some Gαtran/Gαgust-IR cells co-expressed PYY- or CCK-, and few Gαtran/Gαgust-IR cells were positive for GHR-IR. In the cecum, several Gαtran or Gαgust-IR cells were IR for 5-HT. Finally, many Gαtran/Gαgust cells containing 5-HT were observed in the villi and crypts of the cloaca, whereas there were few Gαtran or Gαgust/CCK-IR cells. The demonstration that Gα-subunits are expressed in the chicken GI enteroendocrine system supports the involvement of taste signaling machinery in the chicken chemosensing processes.

INPP4B overexpression and c-KIT downregulation in human achalasia.

BACKGROUND: Achalasia is a rare motility disorder characterized by myenteric neuron and interstitial cells of Cajal (ICC) abnormalities leading to deranged/absent peristalsis and lack of relaxation of the lower esophageal sphincter. The mechanisms contributing to neuronal and ICC changes in achalasia are only partially understood. Our goal was to identify novel molecular features occurring in patients with primary achalasia. METHODS: Esophageal full-thickness biopsies from 42 (22 females; age range: 16-82 years) clinically, radiologically, and manometrically characterized patients with primary achalasia were examined and compared to those obtained from 10 subjects (controls) undergoing surgery for uncomplicated esophageal cancer (or upper stomach disorders). Tissue RNA extracted from biopsies of cases and controls was used for library preparation and sequencing. Data analysis was performed with the "edgeR" option of R-Bioconductor. Data were validated by real-time RT-PCR, western blotting and immunohistochemistry. KEY RESULTS: Quantitative transcriptome evaluation and cluster analysis revealed 111 differentially expressed genes, with a P ≤ 10-3 . Nine genes with a P ≤ 10-4 were further validated. CYR61, CTGF, c-KIT, DUSP5, EGR1 were downregulated, whereas AKAP6 and INPP4B were upregulated in patients vs controls. Compared to controls, immunohistochemical analysis revealed a clear increase in INPP4B, whereas c-KIT immunolabeling resulted downregulated. As INPP4B regulates Akt pathway, we used western blot to show that phospho-Akt was significantly reduced in achalasia patients vs controls. CONCLUSIONS & INFERENCES: The identification of altered gene expression, including INPP4B, a regulator of the Akt pathway, highlights novel signaling pathways involved in the neuronal and ICC changes underlying primary achalasia.

Downregulation of neuronal vasoactive intestinal polypeptide in Parkinson's disease and chronic constipation.

BACKGROUND: Chronic constipation (CC) is a common and severe gastrointestinal complaint in Parkinson's disease (PD), but its pathogenesis remains poorly understood. This study evaluated functionally distinct submucosal neurons in relation to colonic motility and anorectal function in PD patients with constipation (PD/CC) vs both CC and controls. METHODS: Twenty-nine PD/CC and 10 Rome III-defined CC patients were enrolled. Twenty asymptomatic age-sex matched subjects served as controls. Colonic transit time measurement and conventional anorectal manometry were evaluated in PD/CC and CC patients. Colonoscopy was performed in all three groups. Colonic submucosal whole mounts from PD/CC, CC, and controls were processed for immunohistochemistry with antibodies for vasoactive intestinal polypeptide (VIP) and peripheral choline acetyltransferase, markers for functionally distinct submucosal neurons. The mRNA expression of VIP and its receptors were also assessed. KEY RESULTS: Four subgroups of PD/CC patients were identified: delayed colonic transit plus altered anorectal manometry (65%); delayed colonic transit (13%); altered manometric pattern (13%); and no transit and manometric impairment (9%). There were no differences in the number of neurons/ganglion between PD/CC vs CC or vs controls. A reduced number of submucosal neurons containing VIP immunoreactivity was found in PD/CC vs controls (P<.05). VIP, VIPR1, and VIPR2 mRNA expression was significantly reduced in PD/CC vs CC and controls (P<.05). CONCLUSIONS AND INFERENCES: Colonic motor and rectal sensory functions are impaired in most PD/CC patients. These abnormalities are associated with a decreased VIP expression in submucosal neurons. Both sensory-motor abnormalities and neurally mediated motor and secretory mechanisms are likely to contribute to PD/CC pathophysiology.

Localization of peripheral autonomic neurons innervating the boar urinary bladder trigone and neurochemical features of the sympathetic component.

The urinary bladder trigone (UBT) is a limited area through which the majority of vessels and nerve fibers penetrate into the urinary bladder and where nerve fibers and intramural neurons are more concentrated. We localized the extramural post-ganglionic autonomic neurons supplying the porcine UBT by means of retrograde tracing (Fast Blue, FB). Moreover, we investigated the phenotype of sympathetic trunk ganglion (STG) and caudal mesenteric ganglion (CMG) neurons positive to FB (FB+) by coupling retrograde tracing and double-labeling immunofluorescence methods. A mean number of 1845.1±259.3 FB+ neurons were localized bilaterally in the L1-S3 STG, which appeared as small pericarya (465.6±82.7 µm2) mainly localized along an edge of the ganglion. A large number (4287.5±1450.6) of small (476.1±103.9 µm2) FB+ neurons were localized mainly along a border of both CMG. The largest number (4793.3±1990.8) of FB+ neurons was observed in the pelvic plexus (PP), where labeled neurons were often clustered within different microganglia and had smaller soma cross-sectional area (374.9±85.4 µm2). STG and CMG FB+ neurons were immunoreactive (IR) for tyrosine hydroxylase (TH) (66±10.1% and 52.7±8.2%, respectively), dopamine beta-hydroxylase (DßH) (62±6.2% and 52±6.2%, respectively), neuropeptide Y (NPY) (59±8.2% and 65.8±7.3%, respectively), calcitonin-gene-related peptide (CGRP) (24.1±3.3% and 22.1±3.3%, respectively), substance P (SP) (21.6±2.4% and 37.7±7.5%, respectively), vasoactive intestinal polypeptide (VIP) (18.9±2.3% and 35.4±4.4%, respectively), neuronal nitric oxide synthase (nNOS) (15.3±2% and 32.9±7.7%, respectively), vesicular acetylcholine transporter (VAChT) (15±2% and 34.7±4.5%, respectively), leu-enkephalin (LENK) (14.3±7.1% and 25.9±8.9%, respectively), and somatostatin (SOM) (12.4±3% and 31.8±7.3%, respectively). UBT-projecting neurons were also surrounded by VAChT-, CGRP-, LENK-, and nNOS-IR fibers. The possible role of these neurons and fibers in the neural pathways of the UBT is discussed.

Partial transformation from fast to slow muscle fibers induced by deafferentation of capsaicin-sensitive muscle afferents.

Mechanical and histochemical characteristics of the lateral gastrocnemius (LG) muscle of the rat were examined 21 days after capsaicin injection into the LG muscle. The capsaicin caused a decrease in generation rate of twitch and tetanic tension and an increase in fatigue resistance of LG muscle. The histochemical muscle fiber profile evaluated by myosin adenosine triphosphatase and reduced nicotinamide adenine dinucleotide tetrazolium reductase methods showed an increase of type I and IIC fibers and a decrease of the type IIB in whole muscle, and a decrease of the IIA, IIX fibers in the red part accompanied by their increase in the white part. Therefore the capsaicin treatment, which selectively eliminated fibers belonging to the III and IV groups of muscle afferents, induced muscle fiber transformation from fast contracting fatiguing fibers to slowly contracting nonfatiguing ones.

Analysis of the sternotrachealis muscle fibers in some Anseriformes: histochemistry and sex differences.

Histochemical characteristics and sizes of the fibers of the sternotrachealis (ST) muscle have been investigated in some Anseriformes (mallard, Pekin duck, Muscovy duck, and goose) of both sexes. A sexual dimorphism has been shown in the muscle of the species examined. In the mallard and Pekin duck, the male ST muscle shows type IIIA fibers in addition to the type I, IIA, and IIB fibers observed also in the female. In the Muscovy duck, the male muscle has only type I and IIA fibers, whereas the female muscle presents type I fibers and both types IIA and IIB fibers. Moreover, the mean frequencies for each fiber type were significantly different between males and females. In the goose, both male and female muscles present only type I and IIA fibers. In all the species examined, the mean areas of each fiber type are significantly different between male and female, being always larger in the male muscles. The anatomical sexual dimorphism observed in the ST muscle is discussed in relation to function.