Summal"yTo determine the functional role of the two isoforms of Fc3,RIII (CD16) (IliA, IIIB), the signal transduction capabilities of wild-type and mutant forms of these receptors were analyzed in transfected lymphoid, myeloid, and fibroblastic cell lines. Functional reconstitution of receptor signalling was observed in hematopoietic T and mast cells, and was absent in nonhematopoietic (CHO) cells.Fc~RIIIA, a hetero-o[igomeric receptor composed of a ligand-binding subunit c~ and dimeric 3/chains, generated both proximal and distal responses in Jurkat and P815 cells, typical of what is seen in natural killer cells and macrophages upon receptor activation. In contrast, Fc3/RIIIB, which is normally attached to the cell surface via a glycosyl-phosphatidylinositol anchor, was incapable of transducing signals. After crosslinking, Fc3,KIIIA signalling was dependent only upon the 3, chain. Fc3/RIIIA chimeras in which the ol subunit transmembrane and cytoplasmic domains were substituted with the corresponding 3/chain sequences functioned as well as wildtype hetero-oligomeric receptors. These data indicate that the ability of the FcyRIIIA complex to activate the appropriate pathways for cell activation is cell-type restricted and independent of the transmembrane and cytoplasmic domains of the c~ subunit. The presence of the 3/chain is responsible for the assembly of, as well as the signal transduction by, the functional cell surface complex.

5-Hydroxytryptamine (5-HT) is one of the major chemical mediators released in injured and

Functional changes of the enteric nervous system have been observed under inflammatory states of inflammatory bowel disease increasing the endotoxin level. The aim of the present study was to determine the effect of lipopolysaccharides (LPS) on myenteric neuron-glia interaction in vitro. We examined the increase of the intracellular Ca(2+) concentration ([Ca(2+)](i)) and the release of interleukin-1beta (IL-1beta) or prostaglandin E(2) (PGE(2)) and COX-2 expression in myenteric plexus cells from the rat intestine induced by LPS. LPS potentiated BK-induced [Ca(2+)](i) increases in both myenteric neurons and enteric glial cells, which were suppressed by a B1R antagonist. Only in enteric glial cells, a B1R agonist increased [Ca(2+)](i). The effects of LPS were blocked by pretreatment with an interleukin-1 receptor antagonist or by reducing the density of enteric glial cells in culture. LPS prompted the release of IL-1beta from enteric glial cells. The augmenting effects of IL-1beta on the BK-induced neural [Ca(2+)](i) increase and PGE(2) release from enteric glial cells were abolished by a phospholipase A(2) (PLA(2)) inhibitor and a COX inhibitor, and partly suppressed by a COX-2 inhibitor. IL-1beta up-regulated the COX-2 expression in enteric glial cells. LPS promotes IL-1beta secretion from enteric glial cells, resulting in augmentation of the neural response to BK through PGE(2) release via glial PLA(2) and COX-2. The alteration of the regulatory effect of glial cells may be the cause of the changes in neural function in the enteric nervous system in inflammatory bowel disease.

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]i similar to that induced by P2Y agonists. At Ϫ60 mV, ATP evoked a slowly inactivating inward current that was suppressed by the removal of extracellular Na ϩ concentration. The current-voltage relation for ATP showed an inward rectification with the reversal potential of about 0 mV. The apparent rank order of potency for the purinoceptor agonist-induced increases of [Ca 2ϩ ]i was ATP Ն adenosine 5Ј-O-3-triphosphate Ն CTP Ն 2-methylthio-ATP Ͼ benzoylbenzoyl-ATP. A similar potency order was obtained with current responses to these agonists. P2 antagonists inhibited inward currents induced by ATP. Ca 2ϩ and Mg 2ϩ suppressed the ATP-induced current, and Zn 2ϩ , Cu 2ϩ , and protons potentiated it. RT-PCR and immunocytochemical studies showed the expression of P2X 2 receptors in cultured rat myenteric neurons. These results suggest that ATP mainly activates ionotropic P2X 2 receptors, resulting in a [Ca 2ϩ ]i increase dependent on [Ca 2ϩ ]o in rat myenteric neurons. A small part of the ATP-induced [Ca 2ϩ ]i increase may be also mediated via a P2Y receptor-related mechanism. enteric neuron; fura-2; patch clamp; purinoceptor THE ENTERIC NERVOUS SYSTEM (ENS) plays a key role in controlling various gastrointestinal functions, including motor activity, secretion, absorption, and local circulation (60). The ENS is composed of functionally different neurons that contain a variety of potential neurotransmitters and express receptors for these substances (20). Accumulation of ATP in the extracellular space evokes various responses in neurons, immune/inflammatory cells, smooth muscle, and glandular epithelium by activating different P2 purinoceptor subtypes (10, 49). In the gastrointestinal tract, ATP was first recognized as an inhibitory neurotransmitter released from myenteric motor neurons to relax smooth muscles (16). In enteric neurons of the guinea pig, a part of the fast excitatory postsynaptic potentials (fEPSPs) is suppressed by a nicotinic receptor antagonist, and the remaining fEPSPs are further reduced by P2 purinoceptor antagonists (22), indicating the presence of purinergic excitatory neurotransmission in the ENS (45). An experiment using mice lacking the P2X receptor subtype suggested that ATP was involved in fEPSPs (51). Moreover, it has recently been reported that ATP functions as a putative sensory mediator from epithelial sources to the intrinsic sensory nerve terminals (3). Therefore, purinergic signaling in the ENS is considered to be important in the regulation of gastrointestinal functions (17).P2 purinoceptors are identified as ionotropic and metabotropic receptors based on their pharmacological properties, mechanisms of signal transudation, and deduced amino acid sequences (32). Ionotropic P2 purinoceptors, P2X, are ligandgated nonselective cation channels that are permeable for Na ϩ and Ca 2ϩ (44). Seven P2X receptors, identified as P2X 1 -P2X 7 , have been cloned. In the ENS, P2X 2 (11, 58), P2X 3 (48), P2X 2 and P2X 3 (59, 61), and P2X 7 (30) have been detected immunohistochemically. It has...