Extrinsic denervation elevates neuronal aromatic L-amino acid decarboxylase immunoreactivity in rat small intestine

In order to clarify further the neural control of digestive tract function, we have compared the neuronal localization of tyrosine hydroxylase (TH) and aromatic amino acid decarboxylase (AADC) in rat small intestine. Immunoreactivity for TH was found in numerous varicose axons associated with neurons of the enteric plexuses and in axons within the circular muscular coat and the mucosal villi. Axons with AADC immunoreactivity had a similar distribution, but were sparser in the enteric plexuses and musculature than those containing TH. Chronic extrinsic denervation of a segment of intestine removed all TH-positive nerves from that region. By contrast, the intensity of AADC immunoreactivity was enhanced and more AADC-positive axons were visible than in adjacent intact areas of intestine. The AADC-positive axons appear to represent the intrinsic 'amine-handling' neurons rather than intrinsic tryptaminergic neurons or extrinsic dopaminergic neurons, and the effect on AADC activity of removing the extrinsic nerve supply suggests that this normally exerts some restraining influence on the metabolism of the 'amine-handling' population.     

Distribution of aromatic l-amino acid decarboxylase in tissues of skipjack tuna using l-DOPA as the substrate

Aromatic L-amino acid decarboxylase activity was measured in brain, heart, intestine, kidney, liver, muscle, pyloric caeca, spleen and stomach of skipjack, using L-3,4–dihydroxyphenylalanine as the substrate. Aromatic L-amino acid decarboxylase activity was found to be present in all of the organs studied. The highest activity was found in the intestine (1774 μmol min ⁻¹ g⁻¹ wet wt of tissue). The liver showed the lowest activity (48.7 umol min ⁻¹ g ⁻¹).     

Neuronal distribution and subcellular localization of HCNP-like immunoreactivity in rat small intestine

A novel peptide, hippocampal cholinergic neurostimulating peptide (HCNP), originally purified from young rat hippocampus, affects the development of specific cholinergic neurons of the central nervous system in vitro. In this study, HCNP-like-immunoreactive nerve processes and nerve cell bodies were identified by electron microscopic immunocytochemistry in the rat small intestine. Labeled nerve processes were numerous in the circular muscle layer and around the submucosal blood vessels. In the submucosal and myenteric plexuses, some HCNP-like-immunopositive nerve cell bodies and nerve fibers were present. The reaction product was deposited on the membranes of various subcellular organelles, including the rough endoplasmic reticulum, Golgi saccules, ovoid electron-lucent synaptic vesicles in axon terminals associated with submucosal and myenteric plexuses, and the outer membranes of a few mitochondria. The synaptic vesicles of HCNP-like-positive terminals were 60–85 nm in diameter. The present data provide direct immunocytochemical evidence that HCNP-like-positive nerve cell bodies and nerve fibers are present in the submucosal and myenteric plexuses of the rat small intestine. An immunohistochemical light microscopic study using mirror-image sections revealed that in both the submucosal and myenteric ganglia, almost all choline acetyltransferase (ChAT)-immunoreactive neurons were also immunoreactive for HCNP. These observations suggest (i) that HCNP proper and/or HCNP precursor protein is a membrane-associated protein with a widespread subcellular distribution, (ii) that HCNP precursor protein may be biosynthesized within neurons localized in the rat enteric nervous system, and (iii) that HCNP proper and/or HCNP precursor protein are probably stored in axon terminals.     

Micromolar concentration of kynurenic acid in rat small intestine

Kynurenic acid is an antagonist of glutamate and alpha-7 nicotinic acetylcholine receptors and an agonist of the G: -protein-coupled receptor GPR35, which is predominantly expressed in immune and gastrointestinal tissues. In this study, we report that kynurenic acid is present in the lumen of rat small intestine in micromolar concentration sufficient to affect the GPR35 receptor. Moreover, we show that kynurenic acid can be produced by Escherichia coli. We suggest that kynurenic acid may modulate gastrointestinal function and integrity.     

A single gene codes for aromatic L-amino acid decarboxylase in both neuronal and non-neuronal tissues

We have sought to determine whether aromatic L-amino acid decarboxylase which functions as a neurotransmitter biosynthetic enzyme in neuronal cells can be distinguished from an enzyme with similar activity found in peripheral tissues where no neurotransmitters are synthesized. Aromatic L-amino acid decarboxylase was purified to electrophoretic homogeneity from bovine adrenal medulla, and highly specific antibodies were produced. In addition, a DNA clone complementary to aromatic L-amino acid decarboxylase mRNA was isolated by immunological screening of a lambda gt11 cDNA expression library. We have used these antibodies and cDNA probes for biochemical, immunochemical, and molecular analyses. A single form of aromatic L-amino acid decarboxylase is detected in rat and bovine tissue. Specifically, aromatic L-amino acid decarboxylase protein is biochemically and immunochemically indistinguishable in brain, liver, kidney, and adrenal medulla. Hybridization to aromatic L-amino acid decarboxylase cDNA identifies a single mRNA species of 2.3 kilobase pairs in rat tissue. Furthermore, Southern blot analysis reveals that a single gene codes for aromatic L-amino acid decarboxylase.     

Differential stimulation of S-adenosylmethionine decarboxylase by difluoromethylornithine in the rat colon and small intestine.

The effects of sodium cyclobutyrate, a synthetic hydrocholeretic drug, on biliary lipid secretion and on the biliary outputs of several plasma-membrane enzymes were investigated in anaesthetized rats. Administration of a single oral dose of cyclobutyrol (0.72 mmol/kg body wt.) reduced biliary concentration and output of cholesterol and phospholipid. However, bile acid secretion was not significantly modified. This uncoupling effect of lipid secretion remained even when the choleretic response to the drug had ceased. It additionally led to a statistically significant decrease in the cholesterol/bile acid and phospholipid/bile acid molar ratios and in the lithogenic index of the bile. The biliary outputs of the plasma-membrane enzymes alkaline phosphatase and gamma-glutamyltransferase were markedly reduced by the drug. When cyclobutyrol was administered to rats which had been previously fed with a high-cholesterol diet, the effects of cyclobutyrol persisted, but were less marked. Our results demonstrate that the bile acid-independent choleresis induced by cyclobutyrol (related to its pharmacokinetic effect) is accompanied by a pharmacodynamic action that selectively reduces the secretion of biliary lipids. This is due to an uncoupling of the secretion of cholesterol and phospholipids from that of bile acids. Possible explanations for the biliary response to cyclobutyrol are discussed.     

Slow-waves in rat small intestine

Rat small intestine generates rhythmic slow-wave activity. The slow-waves are not eliminated by ouabain application or incubation in potassium free solution. Exposure to low sodium or calcium free solution decreases slow-wave activity. Incubation in sodium and calcium free solution eliminates activity. It is concluded that rat small intestinal slow-waves may not result from the same mechanism as in the cat.     

Calcitonin gene-related peptide-like immunoreactivity in the human small intestine.

Calcitonin-gene-related-peptide (CGRP)-like immunoreactivity was localized in nerve fibres, neuronal somata and in mucosal endocrine cells of the human small intestine. Immunoreactive enteric neurons were more numerous in the submucous plexuses than in the myenteric plexus. Morphologically, they predominantly had the appearance of type II neurons. The majority of the CGRP-like immunoreactive nerve fibres ran within the ganglionic nerve plexuses. Only a small proportion could be observed in the lamina propria, the lamina muscularis mucosae, or the circular and longitudinal outer smooth muscle layer. These findings suggest that within the wall of the human small intestine neuronal CGRP of either extrinsic or intrinsic origin exerts its effect chiefly on other enteric neurons, and might be indirectly involved in the regulatory functions of the human small intestine.     

Absorption of nicotinic acid and nicotinamide from rat small intestine in vitro

Intestinal absorption of nicotinic acid and nicotinamide was studied using everted sacs of rat small intestine. Transport down the concentration gradient showed saturation kinetics at low concentrations and linear kinetics at higher concentrations. Addition of ouabain or omission of sodium ions decreased absorption. Neither compound was absorbed against a concentration gradient. The mode of transport was thought to be carrier-mediated facilitated diffusion at lower concentrations masked by passive diffusion at higher concentrations.     

Chain-shortening of erucic acid and microperoxisomal beta-oxidation in rat small intestine.

The ability of rat small intestine to chain-shorten C22:1 fatty acids was investigated. Radioactive chain-shortened products, mainly C18:1, were demonstrated in intestinal-lymph lipids after intraluminal injection of [14-14C]erucic acid. Chain-elongation to C24:1 was also observed. Adaptation to a diet containing C22:1 fatty acids (partially hydrogenated-marine-oil diet) slightly increased the percentage of chain-shortened products. Microperoxisomal beta-oxidation activity, measured as CN(-)-insensitive palmitoyl-CoA-dependent NAD+ reduction, was detected in a microperoxisome-enriched fraction from mucosal scrapings. This activity was increased 1.9-fold by a soya-bean-oil diet, and 2.7-fold by a diet containing partially hydrogenated marine oil.     

Epitope mapping of human aromatic L-amino acid decarboxylase

Autoimmune polyendocrine syndrome type I (APS I) is a rare hereditary condition considered a model disease for organ specific autoimmunity. A wide range of autoantibodies targeting antigens present in the affected organs have been identified. Autoantibodies against aromatic L-amino acid decarboxylase (AADC) are present in about 50% of APS I patients. In order to increase our understanding of autoantibody specificity in APS I, the aim of the present study was to localize target regions on AADC recognized by sera from APS I patients. Using several complementing strategies, we have shown that autoantibodies against AADC mainly recognize conformational epitopes. The major antigenic determinants were detected N-terminally to amino acid residue 237. Replacement of amino acids 227-230 (ERDK) with alanine residues reduced the reactivity towards AADC by >80% in all patient sera tested, suggesting that amino acids 227-230 are an important part of an immunodominant epitope.     

Selective sympathetic denervation induced by 6-hydroxy-dopamine in the small intestine.

The structure of the nerve fibres in the intact and neurally isolated intestine of the cat was studied after 6-OHDA treatment by fluorescence and electron microscopy. A gradual disappearance of green fluorescence in the monoaminergic fibers was observed. Degeneration of nerve fibres containing 30--60 nm agranular and 30--60 nm densecore (granular) vesicles could be seen in all layers of the small intestine. Sporadically, degeneration of the presynaptic elements occurred also in the synapses of myenteric ganglia. A large number of degenerating fibres could be observed in close relation to blood vessels. These were thought to be monoaminergic because 6-OHDA selectively affected their terminals. In the chronically isolated small intestine degenerated nerve processes could not be observed following 6-OHDA treatment. It is concluded that the intrinsic nerve elements do not contain monoamines. Accordingly, the previously observed yellow fluorescence of the nerve elements of the isolated small intestine might be due to some other kind of neurotransmitter, possibly tryptamine.