On the mechanisms and putative pathways involving neuroimmune interactions

Bidirectional interdependence between the immune system and the CNS involves the intervention of common cofactors. Cytokines are endogenous to the brain, endocrine and immune systems. These shared ligands are used as a chemical language for communication. Such interaction suggests an immunoregulatory role for the brain, and a sensory function for the immune system. Interplay between the immune, nervous and endocrine systems is associated with effects of stress on immunity. Cytokines are thus capable of modulating responses in the CNS, while neuropeptides can exert their effects over cellular groups in the immune system. One way is controlled by the HPA axis, a coordinator of neuroimmune interactions that is essential to unravel in order to elucidate vital communications in a manner that this crosstalk remains a cornerstone in perpetuating a stance of homeostasis.     

The role of glutaminase in the small intestine

Glutaminase is the enzyme which hydrolyses glutamine, the main respiratory fuel of the intestine, to yield glutamate and ammonia. Glutaminase has a central role in intestinal metabolism: the products of the reaction catalyzed by glutaminase can be transaminated, catabolized to yield energy or used for the biosynthesis of pyrimidine nucleotides. Experimental treatments which deprive the intestine of glutamine induce intestinal atrophy. In this review, attention is paid to the role of glutaminase in intestinal metabolism. Background information on the structure, kinetics and distribution of glutaminase precede a discussion of the metabolism of glutamine within the intestine. In closing, we review the factors known to regulate glutaminase activity and emphasise that the regulation of glutaminase within the intestine is poorly understood.     

PGE(2) receptors and their intracellular mechanisms in rabbit small intestine

The effects of PGE(2) on longitudinal smooth muscle, the intracellular mechanisms involved, and the localization of EP receptors were investigated in rabbit small intestine. PGE(2) evoked contractions in small intestine that were reduced by tetrodotoxin and hexamethonium. 17-Phenyl trinor PGE(2), sulprostone, misoprostol and 16,16-dimethyl PGE(2) evoked contractions. Butaprost did not modify spontaneous motility. AH 6809 reduced PGE(2) and 17-phenyl trinor PGE(2)-induced contractions. Verapamil, Ca(2+) free medium, staurosporine, forskolin, theophylline, and rolipram diminished, while IP-20 and H-89 increased PGE(2)-induced contractions. Western blot analysis showed protein bands of 41kDa for EP(1), 71kDa for EP(2) and 62kDa for EP(3) receptors. EP(1), EP(2) and EP(3) receptors were detected in neurons of the myenteric and submucosal ganglia, but only EP(3) receptors were found in smooth muscle layers. This study did not detect EP(4) receptor. PGE(2)-induced contractions would be mediated through EP(1) and EP(3) receptors, and voltage-dependent Ca(2+) channels, protein kinase C, and cAMP would be implicated in these responses.     

The cross-ply arrangement of collagen fibres in the submucosa of the mammalian small intestine

Summary Whole-mount preparations of the submucosa were made from the small intestine of rats, guinea-pigs, rabbits and sheep. In the distended intestine the collagen fibres ran straight and approximately parallel to the serosal surface. They formed a characteristic lattice, with two arrays of fibres running diagonally in a clockwise and an anticlockwise direction, and making an angle of 50°–55° with the longitudinal axis of the intestine. This collagenfibre lattice was flexible and changed with the movements of the intestinal wall; when the radial distension predominated, the angle between collagen fibres of the submucosa and longitudinal axis of the intestine increased to 60°–65°, and when the longitudinal distension predominated the angle decreased to about 30°.     

Analysis of the role of adrenergic receptors in the regulation of small intestine motor activity in sheep

In sheep with chronic fistulae of the small intestine and rumen the participation of alpha- and beta-adrenergic receptors in the regulation of the motor activity of the small intestine was studied by the method of pharmacological analysis. The movements of the fistulated parts of the alimentary tract were recorded by the balloon method. Slow intravenous infusion of isoprenaline inhibited the contractions of the small intestine. This inhibitory effect of isoprenaline was abolished by propranolol. Intravenous phenylephrine inhibited the motor activity of this intestinal part as well. The effect of phenylephrine was abolished by pretreatment with dihydroergotamine. In the small intestine of sheep stimulation of the alpha and beta adrenergic receptors decrease the motor activity of intestine.     

Bacteria-lectin interactions in phytohemagglutinin-induced bacterial overgrowth of the small intestine

The mechanism of phytohemagglutinin-induced bacterial overgrowth of the small bowel in the rat was studied. Interaction of the lectin with bacterial isolates selected at random from those that comprised the major population of the overgrowth was determined. In both bacterial agglutination assays and glycocalyx stabilization, no specific association between lectin and bacteria was seen. In three independent binding assays phytohemagglutinin was not found to increase bacterial adherence to washed intestinal mucosa. Phytohemagglutinin would not appear to act, therefore, as a direct ligand to mediate bacterial adherence or to modify the mucosal surface to increase bacterial adherence.     

Motilin receptors in rabbit stomach and small intestine

Motilin receptors in rabbit antral and duodenal smooth muscle tissue were characterized by direct binding technique using 125I-labeled porcine motilin as a tracer ligand. Binding at 30 degrees C was maximal at 90 min, was saturable and partially reversible. Displacement studies with natural porcine motilin, synthetic leucine-motilin or norleucine-motilin indicated a dissociation constant (Kd) of 1.1 +/- 0.3 nM and a maximal binding capacity (Bmax) of 42 +/- 10 fmol/mg protein. Binding was unaffected by glucagon, pancreatic polypeptide and somatostatin, but substance P interfered via an unknown mechanism. By density gradient centrifugation motilin receptors were shown to be present in plasma membranes. Binding could only be demonstrated in preparations from antrum and upper duodenum. These observations provide evidence for a localized target region for motilin in the gastrointestinal tract, and for a direct interaction of motilin with gastrointestinal smooth muscle tissue.     

The role of stacking interactions in the mechanisms of clonidine binding

The stacking interactions of the clonidine aromatic ring with the aromatic rings of Phe or Tyr of alpha2-adrenoreceptor and Tyr aromatic ring of the pore of the tetradotoxin-resistant channel have been investigated. Ab initio quantum chemical calculations for a model system of two parallel aromatic rings were performed by GAMESS software with 6-31G** basis set in the framework of the Moller-Plesset second-order perturbation theory with full geometry optimization without any symmetry. It was shown that the parallel shifted conformation of two aromatic rings is energetically most favorable. The 2,6-chlorination of one of the benzene rings leads to the amplification of the stacking interaction, an increase in the relative shift of the rings and possible growth of both the hypotensive and analgetic functions of clonidine due to the increase in the binding energy. The 4-fluoridization of the clonidine benzene ring can amplify its analgetic function but practically excludes its hypotensive action.     

The role of cell interactions in the differentiation of teratocarcinoma-derived parietal and visceral endoderm

Cell interactions have been implicated in the differentiation of visceral and parietal endoderm in the developing mouse embryo. Embryoid bodies formed from F9 embryonal carcinoma cells have been useful in characterizing the events which lead to endoderm formation. As part of our effort to specify the interactions which may be involved in this process we have isolated visceral endoderm-like cells (VE) from F9 embryoid bodies and cultured them under various conditions. Using a combination of immunoprecipitation and enzyme-linked immunosorbent assay, we demonstrate that monolayer culture of these cells on a number of different substrates leads to a dramatic decrease in the level of alphafetoprotein (AFP), a VE-specific marker. Northern blot analysis of AFP mRNA indicates very low levels of this message are present after 48 hr in monolayer culture. Coincident with the drop in AFP levels is an increase in the levels of the cytokeratin Endo C and tissue plasminogen activator, both markers for parietal endoderm (PE). Morphological evidence at the ultrastructural level supports a transition from VE to PE. In contrast, the VE phenotype can be maintained in vitro by interaction with aggregates, but not monolayers, of stem cells. In addition, culturing the cells on the curved surface of gelatin-coated dextran beads, but not on a flat gelatin surface facilitates AFP expression and the cells are morphologically intermediate between VE and PE cells. The potential role of junctional complexes and cell shape are discussed.     

Epithelial distribution of neural receptors in the guinea pig small intestine

Neural and paracrine agents, such as dopamine, epinephrine, and histamine, affect intestinal epithelial function, but it is unclear if these agents act on receptors directly at the enterocyte level. The cellular localization and villus-crypt distribution of adrenergic, dopamine, and histamine receptors within the intestinal epithelium is obscure and needs to be identified. Single cell populations of villus or crypt epithelial cells were isolated from the jejunum of adult guinea pigs. Enterocytes were separated from intraepithelial lymphocytes by flow cytometry and specific binding was determined using fluorescent probes. Alpha1-adrenergic receptors were located on villus and crypt intraepithelial lymphocytes and enterocytes. Beta-adrenergic receptors were found on villus and crypt enterocytes. Dopamine receptors were found on all cell types examined, whereas histamine receptors were not detected (<10% for each cell population). These studies demonstrated that (1) receptors for epinephrine and dopamine exist on epithelial cells of the guinea pig jejunum, (2) beta-adrenergic receptors are found primarily on villus and crypt enterocytes and (3) intraepithelial lymphocytes contain alpha1-adrenergic, but have few beta-adrenergic, receptors. The presence of neural receptors suggests that these agents are acting, at least in part, at the enterocyte or intraepithelial lymphocyte levels to modulate intestinal and immune function.     

Cytokines as mediators of neuroimmune interactions

Cytokines regulate numerous physiological and pathological processes in the central nervous system (CNS), i.e. they function both as immune regulators and neuromodulators. Acting upon the CNS via different ways, cytokines, mainly proinflammatory ones IL-1beta and TNF-alpha, can disturb physiological functions of the CNS, cause neurotoxic and neurodegenerative damage and stimulate IL-1beta synthesis in hypothalamus nuclei and posterior pituitary. They can produce stress-like effects upon the CNS and affect the activity of the axis hypothalamus--pituitary--adrenal glands, levels of neuropeptides in hypothalamic regions of brain, synthesis and utilization of central monoamines. These influences can implement the effects of sensitization, which enhances neuroendocrine responses to later stresses. Microglia and astrocytes, secondary messengers and interaction between hypothalamus and anterior pituitary play an important role in range of these processes as well as in the maintenance of Th1/Th2 cytokine balance.     

Neuroimmune interactions in guinea pig stomach and small intestine

Enteric neuroimmune interactions in gastrointestinal hypersensitivity responses involve antigen detection by mast cells, mast cell degranulation, release of chemical mediators, and modulatory actions of the mediators on the enteric nervous system (ENS). Electrophysiological methods were used to investigate electrical and synaptic behavior of neurons in the stomach and small intestine during exposure to beta-lactoglobulin in guinea pigs sensitized to cow's milk. Application of beta-lactoglobulin to sensitized preparations depolarized the membrane potential and increased neuronal excitability in small intestinal neurons but not in gastric neurons. Effects on membrane potential and excitability in the small intestine were suppressed by the mast cell stabilizing drug ketotifen, the histamine H(2) receptor antagonist cimetidine, the cyclooxygenase inhibitor piroxicam, and the 5-lipoxygenase inhibitor caffeic acid. Unlike small intestinal ganglion cells, gastric myenteric neurons did not respond to histamine applied exogenously. Antigenic exposure suppressed noradrenergic inhibitory neurotransmission in the small intestinal submucosal plexus. The histamine H(3) receptor antagonist thioperamide and piroxicam, but not caffeic acid, prevented the allergic suppression of noradrenergic inhibitory neurotransmission. Antigenic stimulation of neuronal excitability and suppression of synaptic transmission occurred only in milk-sensitized animals. Results suggest that signaling between mast cells and the ENS underlies intestinal, but not gastric, anaphylactic responses associated with food allergies. Histamine, prostaglandins, and leukotrienes are paracrine signals in the communication pathway from mast cells to the small intestinal ENS.     

Neurotensin receptors on circular smooth muscle of canine small intestine: role of disulfide bridges

We have studied the effect of sulfhydryl agents on the binding of 125I-tyr3-neurotensin to the purified plasma membranes from circular smooth muscle and on the in vitro response of circular muscle strips of canine small intestine to neurotensin. Dithiothreitol (DTT) enhanced the binding by about 80%. Cysteine (a reductant) also enhanced the binding while cystine (an oxidant) reduced the binding to the similar extent. DTT stimulated the tissue in the organ bath and abolished the stimulatory response to low concentrations of neurotensin. The stimulatory response to acetylcholine was not altered by DTT. The implications of the role of disulfide bridges in the neurotensin response is discussed.     

Endothelin-3 induced mesenteric vasoconstriction and PMN infiltration in the rat small intestine: role of endothelin receptors

The objectives of this study were to characterize endothelin (ET)-3-induced alterations in intestinal hemodynamics and to evaluate whether ET-3 administration alters the tissue levels of polymorphonuclear leukocytes (PMNs) and modulates the epithelial barrier function of the small intestine. ET-3 (100 pmol/kg/min) was infused into the superior mesenteric artery (SMA) for 10 min, and tissue samples were obtained 30 min after terminating the infusion. SMA blood flow was significantly decreased throughout the experiment following ET-3 infusion. Pretreatment with bosentan (ET-A and ET-B receptor antagonist), ET-B receptor antagonist BQ-788 or ET-A receptor antagonist BQ-485 completely inhibited the ET-3-induced decrease in the SMA blood flow. Similar results were obtained from the resistance data, in which ET-3-induced increases in SMA resistance were significantly reduced by all ET receptor antagonists. ET-3 administration significantly elevated tissue MPO activity, blood-to-lumen clearance of (51)Cr-EDTA and caused a marked microscopic damage in the intestinal mucosa. ET-3-induced elevations in tissue PMN infiltration and mucosal damage were significantly inhibited by pretreatments with ET-A or ET-B receptor antagonists. Overall, our data indicate that ET-3 causes microscopic damage, PMN infiltration and mucosal dysfunction in the rat small intestine. In addition, ET-3-induced hemodynamic alterations as well as tissue PMN infiltration and mucosal damage are mediated by both ET-A and ET-B receptors.