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J R Grider

Publications and source records attributed to J R Grider.

At least 73 records · Page 4Linked to original sources
PubMed1990

Identification of neurotransmitters by selective protection of postjunctional receptors.

The neurotransmitter responsible for relaxation of gastric smooth muscle was identified by a technique involving the use of selective ligands to protect postjunctional receptors combined with inactivation of all other receptors with N-ethylmaleimide (NEM). Relaxation in response to field stimulation (80 V, 1 ms, 0.5-8 Hz) and to maximally effective concentrations of vasoactive intestinal peptide (VIP; 1 microM), ATP (1 mM), isoproterenol (1 mM), dibutyryl adenosine 3',5'-cyclic monophosphate (cAMP; 1 mM), and forskolin (1 microM) was measured in muscle strips from guinea pig gastric fundus before and after treatment with NEM (5 microM) for 1 h. Protection of VIP receptors with VIP or the VIP antagonist VIP10-28 fully protected relaxation induced by VIP and by field stimulation, but did not protect relaxation induced by ATP or isoproterenol. Protection of ATP receptors with ATP protected only the response to ATP, but did not protect the response to field stimulation, VIP, or isoproterenol. Relaxation induced by either forskolin or dibutyryl cAMP was not altered by treatment with NEM alone or in the presence of protective ligands, indicating that at the concentrations used NEM inactivated membrane receptors without affecting intracellular relaxation mechanisms. These studies indicate that VIP but not ATP is the neurotransmitter responsible for neurally induced relaxation in the gastric fundus.

Adenosine Triphosphate↗
PubMed1990

Distinct receptors for cholecystokinin and gastrin on muscle cells of stomach and gallbladder.

A technique of receptor protection that enables selective preservation of one receptor type was used in conjunction with preferential antagonists of cholecystokinin (L364,718) and gastrin (L365,260) to characterize cholecystokinin (CCK) and gastrin receptors on isolated single muscle cells of the guinea pig stomach and gallbladder. In gastric muscle cells, CCK-8 and gastrin-17 were equipotent and the concentration causing 50% inhibition of response (IC50) values of both antagonists were independent of the agonist, whereas in gallbladder muscle cells, CCK-8 was more potent than gastrin-17 and the IC50 values of the antagonists were dependent on the agonist. In gastric muscle cells, CCK-8, gastrin-17, and L364,718 protected completely the response to CCK-8 or gastrin-17, and L365,260 protected partially but equally the response to both agonists. In gallbladder muscle cells, CCK-8 and L364,718 protected completely and gastrin-17 and L365,260 protected partially the response to CCK-8; conversely, gastrin-17 and L365,260 protected completely and CCK-8 and L364,718 protected partially the response to gastrin-17. The patterns of response in muscle cells from the two regions indicate the presence of one receptor type in gastric muscle cells that interacts equally with gastrin and CCK, and of two receptor types in gallbladder muscle cells, one of which interacts preferentially with CCK and the other preferentially with gastrin.

Animals↗
PubMed1990

Vasoactive intestinal peptide (VIP) as transmitter of inhibitory motor neurons of the gut: evidence from the use of selective VIP antagonists and VIP antiserum.

The role of vasoactive intestinal peptide (VIP) in neurally mediated relaxation of guinea pig and rat intestine was examined with three putative VIP antagonists: a C-terminal sequence of VIP (VIP10-28), substituted analogs of VIP ([4-Cl-D-Phe6, Leu17]VIP) and growth hormone releasing factor (GRF) ([Ac-Tyr1, D-Phe2] GRF1-29). The three agents inhibited selectively relaxation induced by VIP and its homolog, peptide histidine isoleucine. Inhibition of VIP-induced relaxation in tenia coli and gastric fundic strips was consistent with competitive antagonism. Relaxation induced by field stimulation (80 V; 1 msec; 0.1-16 Hz) which is accompanied by a stoichiometric increase in VIP release was inhibited by the three antagonists in the following order of potency: VIP10-28 greater than VIP analog greater than GRF analog. The descending relaxation component of the peristaltic reflex induced by graded stretch of the orad end of a rat colonic segment which is also accompanied by an increase in VIP release, was inhibited by the three VIP antagonists in the same order of potency: VIP10-28 greater than VIP analog greater than GRF analog. The most potent antagonist, VIP10-28, abolished descending relaxation at the lowest grades of stretch (2 and 4 g) and inhibited relaxation at the highest grades of stretch (10 g) by 79%. The results indicate that VIP (and peptide histidine isoleucine) is the transmitter responsible for neurally induced gastric and intestinal relaxation.

Animals↗
PubMed1989

Identification of neurotransmitters regulating intestinal peristaltic reflex in humans.

The components of the intestinal peristaltic reflex in humans were examined and the neurotransmitters responsible for them identified for the first time i isolated flat sheet segments of intestine. Increasing radial stretch to the caudad end elicited increasing ascending contraction only, whereas increasing radial stretch to the orad end elicited increasing descending relaxation only. Both components were abolished by hexamethonium, implying the participation of cholinergic interneurons in each component. Atropine inhibited ascending contraction only, abolishing the response to low grades of stretch and partially inhibiting the response to high grades of stretch (69% +/- 17%, p less than 0.01). The substance P antagonist [D-Pro2, D-Trp7,9] substance P partially inhibited ascending contraction induced by high grades of stretch only (40% +/- 12%, p less than 0.02). The vasoactive intestinal peptide antagonist [4-Cl-D-Phe6, Leu17]vasoactive intestinal peptide inhibited descending relaxation, abolishing the response to low grades of stretch and partially inhibiting the response to high grades of stretch (40% +/- 4%, p less than 0.001). Release of vasoactive intestinal peptide increased significantly by 91% during descending relaxation only, whereas release of both substance P and substance K increased significantly by 107% during ascending contraction only, supporting the participation of vasoactive intestinal peptide motor neurons in descending relaxation and tachykinin motor neurons as well as cholinergic motor neurons in ascending contraction. The components of the human peristaltic reflex and transmitters regulating them were identical to those found in rat and guinea pig intestine.

Atropine↗
PubMed1989

Somatostatin release from isolated ganglia of the myenteric plexus.

Somatostatin neurons of the myenteric plexus project caudad exclusively within the plexus synapsing with neurons in the same or other ganglia. Isolated ganglia offer a unique opportunity to examine peptide transmitter release from these interneurons. Ganglia were isolated from the myenteric plexus by sequential enzymatic digestion, centrifugation, and filtration. Single ganglia were harvested by suction and dispersed in polyacrylamide gel. The ganglia were placed in chambers (200 ganglia/chamber) and perfused with Krebs medium at the rate of 1 ml/min. Addition of the nicotinic agonist, dimethylphenylpiperazinium (DMPP), caused a concomitant increase in somatostatin (152%; P less than 0.001) and vasoactive intestinal peptide (VIP) (79%; P less than 0.05) release above basal level that was abolished by hexamethonium. Addition of the VIP antagonist, VIP-(10-28), (5 microM), to the perfusate augmented significantly DMPP-induced somatostatin release (440%; P less than 0.05), implying that the concomitant release of VIP attenuates somatostatin release. Addition of VIP (1 microM) to the perfusate abolished somatostatin release, confirming the ability of VIP to inhibit somatostatin release. The study shows that VIP interneurons exert a regulatory feedback control over somatostatin interneurons in the myenteric plexus.

Animals↗
PubMed1989

Tachykinins as transmitters of ascending contractile component of the peristaltic reflex.

The role of tachykinin neurons of the myenteric plexus in the regulation of the intestinal peristaltic reflex was examined in rat colonic segments that enable separate measurement of ascending contraction and descending relaxation. Release of immunoreactive substance P (SP) and substance K (SK) increased (68% for SP; 70% for SK) during ascending contraction and decreased (37% for SP; 13% for SK) during descending relaxation. Exogenous SP and SK augmented ascending contraction and inhibited descending relaxation. Neither SP antiserum nor SK antiserum had any effect on descending relaxation. The antisera had no effect also on ascending contraction induced by low grades of stretch (2 g), but they inhibited significantly contraction induced by high grades (6-10 g) of stretch (52 +/- 8% inhibition with SP antiserum and 48 +/- 8% with SK antiserum). A combination of both antisera abolished ascending contraction induced by high grades of stretch. Atropine inhibited only partially ascending contraction at high grades of stretch. The pattern of inhibition by both antisera and by atropine implied that ascending contraction induced by high grades of stretch was mediated by tachykinin neurons acting directly on smooth muscle cells as well as indirectly via release of acetylcholine from cholinergic motor neurons.

Animals↗
PubMed1988

Characterization of opioid receptors on isolated canine gallbladder smooth muscle cells.

Smooth muscle cells were isolated from the fundus of the canine gallbladder and examined for the presence of opioid receptors. The cells contracted in a concentration-dependent manner in response to three opioid peptides (Met-enkephalin, dynorphin1-13 and Leu-enkephalin), which are known derivatives of opioid precursors present in myenteric neurons of the gut. The order of potency was Met-enkephalin greater than dynorphin1-13 greater than Leu-enkephalin. The contractile response to opioid agonists was selectively inhibited by opioid antagonists (naloxone and Mr2266) but not by muscarinic, CCK/gastrin or tachykinin antagonists. Equivalent responses to the three opioid peptides exhibited differential sensitivity to preferential antagonists of mu (naloxone) and kappa (Mr2266) opioid receptors consistent with the presence of the three main types of opioid receptors (mu, delta and kappa) on canine gallbladder muscle cells.

Animals↗
PubMed1988

Identification of bombesin receptors on isolated muscle cells from human intestine.

Smooth muscle cells were isolated separately from the longitudinal and circular muscle layers of human jejunum obtained at surgery and used to determine whether amphibian bombesin-14 and 3 mammalian homologues, GRP-(1-27), GRP-(18-27) and neuromedin B, can cause contraction by acting directly on muscle cells. Circular and longitudinal muscle cells contracted identically in response to bombesin-14 (C50 2 x 10(-12) M). The contractile response was not affected by selective muscarinic, opioid, CCK or serotonin antagonists but was inhibited by the substance P (SP) derivative, [D-Arg1, D-Pro2, D-Trp7,9, Leu11]SP. All 3 mammalian bombesins were less potent than bombesin-14. GRP-(1-27) and GRP-(18-27) were equipotent (C50 4 x 10(-11) M) but 20 times less potent than bombesin-14. Neuromedin B (C50 6 x 10(-12) M) was 3 times less potent than bombesin-14. All bombesins, however, were more potent than other enteric neuropeptides (e.g., tachykinins, opioid peptides). The study demonstrates conclusively the ability of bombesins to cause direct contraction of intestinal smooth muscle cells.

Bombesin↗
PubMed1988

Functional gradients in muscle cells isolated from gallbladder, cystic duct, and common bile duct.

The existence of a gradient inherent to muscle cells of the biliary tract was examined in muscle cells isolated separately from the fundus of the gallbladder, cystic duct, and common bile duct of the dog. Muscle cells, measured in suspension or as single perfused cells, exhibited a proximal-to-distal gradient expressed by the magnitude of response and the sensitivity to hormonal cholecystokinin octapeptide and neural (acetylcholine and methionine-enkephalin) contractile agonists. Measurements in suspensions showed that cells from the fundus 1) were 7-40 times more sensitive to contractile agonists than cells from the cystic duct and 13-200 times more sensitive than cells from the common bile duct and 2) generated greater maximal contraction. The latter was expressed by the ratio of maximal responses (fundus: cystic duct cells, 1.90 +/- 0.12, P less than 0.001; fundus: common bile duct cells, 1.50 +/- 0.07 P less than 0.001), which was independent of sensitivity to agonists. Similar results were obtained in measurements on single cells with respect to relative sensitivity and to ratio of maximal responses (fundus: cystic duct cells, 1.80 +/- 0.08, P less than 0.001; fundus: common bile duct cells, 1.49 +/- 0.06, P less than 0.001). The ratio of responses to low concentrations of agonists was even higher (three- to fourfold), reflecting both the greater sensitivity and the greater contraction of muscle cells of the fundus. We conclude that a proximal-to-distal biliary gradient exists that is an inherent property of muscle cells from various regions of the biliary tract; the gradient would act to facilitate gallbladder emptying in response to hormonal and neural stimulation.

Acetylcholine↗
PubMed1988

Identification of separate bombesin and substance P receptors on isolated muscle cells from canine gallbladder.

Smooth muscle cells were isolated from the dog gallbladder and used to examine whether bombesin-14 and substance P cause direct contraction and whether this effect is mediated by the same or different receptors. Both peptides caused concentration-dependent contraction that was inhibited by two substance P derivatives, [D-Pro2,D-Trp7,9]SP ([PTT]SP) and [D-Arg1,D-Pro2,D-Trp7,9,Leu11]SP ([APTTL]SP). In nonmuscle cells, these substance P derivatives exhibit preferential antagonism of responses to substance P and bombesin, respectively; accordingly, these properties were used to demonstrate the presence of separate bombesin and substance P receptors. Estimates of the apparent Ki showed that [PTT]SP was 40 times more potent than [APTTL]SP as an antagonist of the response to substance P, whereas [APTTL]SP was 85 times more potent than [PTT]SP as an antagonist of the response to bombesin-14. Estimates of median inhibitory concentration obtained from inhibition curves for the effect of various concentrations of each antagonist on the responses to substance P and bombesin-14 confirmed that [PTT]SP was a preferential antagonist of the response to substance P and [APTTL]SP, a preferential antagonist of the response to bombesin-14. The results demonstrate the presence of separate bombesin and substance P receptors mediating contraction of smooth muscle cells of the dog gallbladder.

Animals↗
PubMed1988

Contraction mediated by Ca++ release in circular and Ca++ influx in longitudinal intestinal muscle cells.

The source of Ca++ responsible for contraction was examined in muscle cells isolated separately from the circular and longitudinal muscle layers of guinea pig and human intestine. Contraction was measured by scanning micrometry and cytosolic-free Ca++ ([Ca++]i) with the fluorescent indicator, quin2. In both species, contraction induced in circular muscle cells by cholecystokinin-8 (CCK-8) and acetylcholine was not affected by withdrawal of Ca++ from the medium or addition of the Ca++ channel blocker, methoxyverapamil, whereas contraction induced by both agonists in longitudinal muscle cells and by depolarizing concentrations of K+ in both cell types was abolished. Depletion of intracellular Ca++ stores with caffeine in Ca++-free medium abolished the response in circular muscle cells. Readdition of Ca++ to the medium for 30 sec restored the response in longitudinal but not circular muscle cells. [Ca++]i, measured in guinea pig muscle cells, increased 3- to 4-fold above resting levels (circular, 70.8 +/- 8.1 nM; longitudinal, 77.4 +/- 9.7 nM) in response to all three contractile agents. The increase in [Ca++]i induced by CCK-8 and acetylcholine in circular muscle cells was not affected by withdrawal of Ca++ from the medium or addition of methoxyverapamil, whereas the response to both agonists in longitudinal muscle cells and to 20 mM K+ in both cell types was abolished. It was concluded that cells from adjacent muscle layers of the intestine mobilize Ca++ differently during agonist-induced contraction, i.e., by Ca++ release in circular and Ca++ influx in longitudinal muscle cells.

Animals↗
PubMed1987

Identification of muscarinic M2 receptors on single muscle cells of the human and guinea pig intestine.

Muscle cells were isolated from the longitudinal muscle layer of guinea pig and human jejunum and used to identify the muscarinic receptor subtype (M1 or M2) that mediates contraction. Single muscle cells were anchored to the ceiling of a minichamber and their contraction was measured in response to acetylcholine, alone and in combination with three muscarinic antagonists: atropine, 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP), and pirenzepine. Estimates of the inhibitory dissociation constants (Ki) were closely similar in human and guinea pig muscle cells (atropine 2.5-5 X 10(-11) M, 4-DAMP 1.9-2.9 X 10(10) M, and pirenzepine 8.2-9.5 X 10(-8) M). Thus, pirenzepine, a preferential M1 antagonist, was 1900-3280 times less potent than atropine and 279-500 times less potent than 4-DAMP. Comparative measurements on longitudinal muscle strips from guinea pig jejunum confirmed the greater potency of atropine and 4-DAMP relative to pirenzepine. Inactivation of muscarinic receptors on single muscle cells with dibenamine showed that only a small fraction of receptors was responsible for the response to acetylcholine. It was concluded that intestinal muscle cells contain a large reservoir of muscarinic M2 receptors that exhibit considerable spareness and heterogeneity.

Acetylcholine↗
PubMed1987

Regional and cellular heterogeneity of cholecystokinin receptors mediating muscle contraction in the gut.

The contractile action of cholecystokinin (CCK) on smooth muscle of the gut is either direct (gallbladder and gastric fundus) or both direct and neurally mediated (small intestine). These regional differences were used to characterize pharmacologically CCK receptors on smooth muscle cells and neurons of the gastric fundus, gallbladder, and ileum of the guinea pig. In circular and longitudinal ileal smooth muscle, tetrodotoxin was used to separate direct from neurally mediated contractile effects. Cholecystokinin receptors on smooth muscle cells were found in all locations. The muscle cells displayed a decreasing order of sensitivity to the C-terminal octapeptide of cholecystokinin as expressed in the median doses, and to the selective cholecystokinin antagonist, proglumide, as expressed in the inhibitory dissociation constants. The median doses of the octapeptide of cholecystokinin ranged from 5.5 nM in gallbladder muscle to 185 nM in circular ileal muscle; the corresponding inhibitory dissociation constants of proglumide ranged from 180 to 437 microM [corrected]. Cholecystokinin receptors on cholinergic neurons were confined to circular and longitudinal ileal muscle; the neurons were 80-300 times more sensitive to the octapeptide of cholecystokinin (D50's 0.5 and 2.3 nM) than the corresponding muscle cells, and 19-21 times more sensitive to proglumide (inhibitory dissociation constants, 20 microM [corrected]). The results provide clear evidence of cellular heterogeneity of cholecystokinin receptors (i.e., difference in sensitivity between muscle cells and neurons from the same location) as well as regional heterogeneity (i.e., difference in sensitivity between muscle cells from various locations).

Animals↗
PubMed1987

Prejunctional inhibition of vasoactive intestinal peptide release.

The role of vasoactive intestinal peptide (VIP) and its homologue, peptide histidine isoleucine (PHI), as neurotransmitters of inhibitory motor nerves of the gut, were examined in strips of guinea pig taenia coli and gastric fundic muscle. The stoichiometry of VIP release and muscle relaxation was determined in the presence and absence of the bee venom peptide, apamin, and the existence of prejunctional VIP/PHI receptors capable of regulating VIP/PHI release was explored. In both types of muscle, relaxation induced by field stimulation was proportional to the amount of VIP released. Apamin inhibited relaxation and VIP release in a dose-dependent manner: maximal relaxation was inhibited by 85-96% at 10(-7)-10(-6) M apamin. Analysis of residual responses showed that apamin did not affect the stoichiometry of VIP release and muscle relaxation. Because apamin had no effect on basal tone or on relaxation induced by exogenous VIP, its effect on neurally induced relaxation was attributed to inhibition of VIP release. Both secretin and PHI inhibited neurally induced VIP release in the two types of muscle. At the optimal concentration of 10(-7) M, secretin inhibited VIP release by 52%, whereas the closer neural homologue, PHI, abolished VIP release. The dose-dependent inhibition of VIP release by PHI, which is cosynthesized and coreleased with VIP, indicates the existence of prejunctional inhibitory VIP/PHI autoreceptors capable of regulating VIP/PHI release.

Animals↗
PubMed1987

Role of opioid neurons in the regulation of intestinal peristalsis.

The participation of opioid neurons in the regulation of peristalsis was examined in a rat colonic segment that permits separate characterization of the components of the peristaltic reflex (ascending contraction and descending relaxation). Naloxone increased descending relaxation and decreased ascending contraction; opioid peptides [methionine-enkephalin (Met-Enk), dynorphin-13, and morphiceptin] had opposite effects. Naloxone increased, and Met-Enk decreased, vasoactive intestinal peptide (VIP) release during each component of the reflex. The changes in VIP release reinforced the direct effects of naloxone and opioid peptides on circular muscle tone, providing an explanation for the effects of these agents on the two components of the peristaltic reflex. Dynorphin release decreased during descending relaxation and increased during ascending contraction, reflecting corresponding changes in opioid neural activity. Based on these results a model is proposed, according to which a decrease in opioid neural activity during the initial phase (i.e., descending relaxation) results in direct and VIP-mediated decrease in circular muscle tone. Restoration of opioid neural activity during the subsequent phase (i.e., ascending contraction) increases circular muscle tone and reinforces the action of tachykinin and cholinergic motor neurons, which are the direct mediators of ascending contraction.

Animals↗
PubMed1987

Role of somatostatin neurons in intestinal peristalsis: facilitatory interneurons in descending pathways.

The role of somatostatin neurons in the regulation of peristalsis was examined in segments of rat colon that permit separate characterization of the ascending contraction and descending relaxation components of the peristaltic reflex. Release of somatostatin and vasoactive intestinal peptide (VIP) increased significantly only during descending relaxation. Preincubation of the segment with somatostatin antiserum (final concentration 1:40) decreased VIP release and descending relaxation. Addition of somatostatin (1 nM to 1 microM) augmented VIP release and descending relaxation in a concentration-dependent manner. Together the results implied that the increase in somatostatin release was coupled to, and responsible for, the increase in VIP release, which in turn was responsible for descending relaxation. The results are consistent with the topography of myenteric VIP neurons (which project into circular muscle) and somatostatin neurons (which project caudad within the plexus) and the pharmacological properties of the two peptides. Somatostatin antiserum had no effect on basal VIP release or ascending contraction, indicating that somatostatin neurons were not involved in the regulation of ascending contraction. The study suggests that somatostatin neurons of the myenteric plexus act as facilitatory interneurons in descending pathways.

Animals↗
PubMed1987

Receptors for substance P on isolated intestinal smooth muscle cells of the guinea pig.

Two radioligands, 125I-labeled substance P (125I-SP) and 125I-labeled substance K (125I-SK), were used to characterize the kinetics and stoichiometry of binding of mammalian tachykinins [substance P (SP), substance K (SK), and neuromedin K (NK)] to smooth muscle cells isolated from the longitudinal muscle layer of guinea pig intestine. Specific binding of 125I-SP and 125I-SK was rapid, saturable, reversible, and temperature dependent. Binding attained 63-70% of steady-state binding within 1 min, coincidentally with the time of optimal contraction. The order of potency with which mammalian tachykinins and the SP antagonist, [D-Pro2, D-Trp7,9]SP, inhibited the binding of both radioligands was identical: SP greater than SK greater than NK greater than [D-Pro2, D-Trp7,9]SP, implying preferential interaction with a site that had highest affinity for SP. SK was 2-3 times, NK 3-4 times, and [D-Pro2, D-Trp7,9]SP 7-23 times less potent than SP (IC50 0.36 nM). Except for NK, the order of potency was similar to that for contraction of isolated muscle cells. The existence of binding sites with even higher affinity was suggested by the ability of muscle cells to contract in response to concentrations as low as 10(-13) M. These binding sites were not detectable at the concentration of radioligands used. It was concluded that a SP receptor is the only tachykinin receptor subtype present on intestinal muscle cells of the guinea pig.

Animals↗