Modulation of hypoglossal motoneuron excitability by NK1 receptor activation in neonatal mice in vitro.
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1. The effects of substance P (SP), acting at NK1 receptors, on the excitability and inspiratory activity of hypoglossal (XII) motoneurons (MNs) were investigated using rhythmically active medullary-slice preparations from neonatal mice (postnatal day 0-3). 2. Local application of the NK1 agonist [SAR(9),Met (O(2))(11)]-SP (SP(NK1)) produced a dose-dependent, spantide- (a non-specific NK receptor antagonist) and GR82334-(an NK1 antagonist) sensitive increase in inspiratory burst amplitude recorded from XII nerves. 3. Under current clamp, SP(NK1) significantly depolarized XII MNs, potentiated repetitive firing responses to injected currents and produced a leftward shift in the firing frequency-current relationships without affecting slope. 4. Under voltage clamp, SP(NK1) evoked an inward current and increased input resistance, but had no effect on inspiratory synaptic currents. SP(NK1) currents persisted in the presence of TTX, were GR82334 sensitive, were reduced with hyperpolarization and reversed near the expected E(K). 5. Effects of the alpha(1)-noradrenergic receptor agonist phenylephrine (PE) on repetitive firing behaviour were virtually identical to those of SP(NK1). Moreover, SP(NK1) currents were completely occluded by PE, suggesting that common intracellular pathways mediate the actions of NK1 and alpha(1)-noradrenergic receptors. In spite of the similar actions of SP(NK1) and PE on XII MN responses to somally injected current, alpha(1)-noradrenergic receptor activation potentiated inspiratory synaptic currents and was more than twice as effective in potentiating XII nerve inspiratory burst amplitude. 6. GR82334 reduced XII nerve inspiratory burst amplitude and generated a small outward current in XII MNs. These observations, together with the first immunohistochemical evidence in the newborn for SP immunopositive terminals in the vicinity of SP(NK1)-sensitive inspiratory XII MNs, support the endogenous modulation of XII MN excitability by SP. 7. In contrast to phrenic MNs (Ptak et al. 2000), blocking NMDA receptors with AP5 had no effect on the modulation of XII nerve activity by SP(NK1). 8. In conclusion, SP(NK1) modulates XII motoneuron responses to inspiratory drive primarily through inhibition of a resting, postsynaptic K+ leak conductance. The results establish the functional significance of SP in controlling upper airway tone during early postnatal life and indicate differential modulation of motoneurons controlling airway and pump muscles by SP. (+info)
Nociception and allodynia/hyperalgesia induced by intrathecal administration of fenvalerate.
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The intrathecal injection of fenvalerate, a sodium channel activator, at doses of 0.01 to 3 microg, dose-dependently induced the duration of a characteristic behavioral syndrome mainly consisting of reciprocal hind limb scratching directed towards caudal parts of the body and biting or licking of the hind legs in mice. Fenvalerate-induced behavior was inhibited by morphine (1-10 mg/kg, i.p.). The characteristic behavior was also inhibited by mexiletine, a sodium channel blocker; MK-801, a N-methyl-D-aspartate ion-channel blocker; and GR82334, a neurokinin-1-receptor antagonist. Calphostin C (3 pmol, i.t.), a protein kinase C inhibitor, inhibited fenvalerate-induced behavior. On the other hand, phorbol-12, 13-dibutyrate (50 pmol, i.t.), a protein kinase C activator, markedly enhanced the fenvalerate-induced behavior. The present results also showed that fenvalerate produced thermal allodynia and hyperalgesia in the tail-flick test. Furthermore, fenvalerate-induced thermal allodynia and hyperalgesia were inhibited by the pretreatment with calphostin C. These results suggest that the intrathecal administration of fenvalerate induces a marked nociceptive response and thermal allodynia/hyperalgesia, and they suggest that tetrodotoxin-resistant sodium channels may play an important role in this effect. (+info)
Effect of tachykinins on ascending and descending reflex pathway in rat small intestine.
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AIM: To examine the effect of tachykinins on the ascending reflex pathway in rat small intestine, we used different selective neurokinin (NK) receptor antagonists (RA): a) NK1-RA: GR-82334 and CP-96.345, b) NK2-RA: MEN-10.376 and L-659.877. The aim was further to investigate the effect of substance P (SP) on the ascending excitatory and descending inhibitory reflex pathway. METHODS: The whole segments of rat ileum (10 cm in length) were studied in an organ bath. Ascending contraction of circle muscle was elicited by anal electrical stimulation (3 Hz, 1 ms, 20 V) and measured as change of intraluminal pressure by a perfused manometric system 2 cm and 4 cm orad of the stimulation site. RESULTS: GR-82334 and CP-96.345 (NK1-RA) caused a significant dose-related inhibition of the oral contraction at a distance of 4 cm: GR-82334 [area: -10 % +/- 8 % (10 nmol/L); -29 % +/ -10 % (1000 nmol/L). P < 0.05, n = 10], CP-96.345 [area: -2 %+/- 6 %(0.1 nmol/L); -14 % +/- 10 % (10 nmol/L). P < 0.01, n = 8], whereas the contractile response at a distance of 2 cm was unaltered (n = 8). In contrast, MEN-10.376 and L-659.877 (NK2-RA) did not alter the amplitude or the area under the curve (n = 10). Neither the NK1- nor the NK2-receptor antagonists had a significant effect on the latency of the reflex response. SP showed a significant increase in the ascending contraction and the descending relaxation (n = 6, P < 0.01). CONCLUSION: These results demonstrate that blockade of NK1-receptors decreases the oral reflex response. Latency of the reflex response remains unchanged, indicating that the effect is not due to an action on interneurons. NK2-receptors do not take part in the ascending reflex in rat small intestine. SP increases the descending relaxant reflex response and ascending contraction. (+info)
Substance P modulates localized calcium transients and membrane current responses in murine colonic myocytes.
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1. Neurokinins contribute to the neural regulation of gastrointestinal (GI) smooth muscles. We studied responses of murine colonic smooth muscle cells to substance P (SP) and NK(1) and NK(2) agonists using confocal microscopy and the patch clamp technique. 2. Colonic myocytes generated localized Ca(2+) transients that were coupled to spontaneous transient outward currents (STOCs). SP (10(-10) M) increased Ca(2+) transients and STOCs. Higher concentrations of SP (10(-6) M) increased basal Ca(2+) and inhibited Ca(2+) transients and STOCs. 3. Effects of SP were due to increased Ca(2+) entry via L-type Ca(2+) channels, and were mediated by protein kinase C (PKC). Nifedipine (10(-6) M) and the PKC inhibitor, GF 109203X (10(-6) M) reduced L-type Ca(2+) current and blocked the effects of SP. 4. SP responses depended upon parallel stimulation of NK(1) and NK(2) receptors. NK(1) agonist ([Sar(9),Met(O(2))(11)]-substance P; SSP) and NK(2) agonists (neurokinin A (NKA) or GR-64349) did not mimic the effects of SP alone, but NK(1) and NK(2) agonists were effective when added in combination (10(-10)-10(-6) M). Consistent with this, either an NK(1)-specific antagonist (GR-82334; 10(-7) M) or an NK(2)-specific antagonist (MEN 10,627; 10(-7) M) blocked responses to SP (10(-6) M). 5. Ryanodine (10(-5) M) blocked the increase in Ca(2+) transients and STOCs in response to SP (10(-10) M). 6. Our findings show that low concentrations of SP, via PKC-dependent enhancement of L-type Ca(2+) current and recruitment of ryanodine receptors, stimulate Ca(2+) transients. At higher concentrations of SP (10(-6) M), basal Ca(2+) increases and spontaneous Ca(2+) transients and STOCs are inhibited. (+info)
Genetic locus half baked is necessary for morphogenesis of the ectoderm.
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The zebrafish epiboly mutants partially block epiboly, the vegetalward movement of the blastoderm around the giant yolk cell. Here, we show that the epiboly mutations are located near the centromere of Linkage Group 7 in a single locus, termed the half baked locus. Nevertheless, except for the similar mutants lawine and avalanche, we find the epiboly traits of each of the alleles to be distinguishable, forming an allelic series. Using in situ analysis, we show that the specification and the formation of the germ layers is unaffected. However, during early gastrulation, convergence movements are slowed in homozygous and zygotic maternal dominant (ZMD) heterozygous mutants, especially in the epiblast layer of the blastoderm. Using triple-mutant analysis with squint and cyclops, we show that ablating involution and hypoblast formation in hab has no effect on the epiboly phenotype on the ventral and lateral sides of the embryo, suggesting that the hypoblast has no role in epiboly. Moreover, the triple mutant enhances the depletion of cells on the dorsal side of the embryo, consistent with the idea that convergence movements are defective. Double-mutant analysis with one-eyed pinhead reveals that hab is necessary in the ectodermal portion of the hatching gland. In ZMD heterozygotes, in addition to the slowing of epiboly, morphogenesis of the neural tube is abnormal, with gaps forming in the midline during segmentation stages; later, ectopic rows of neurons form in the widened spinal cord and hindbrain. Cell transplantation reveals that half baked acts both autonomously and nonautonomously in interactions among cells of the forming neural tube. Together, these results suggest that half baked is necessary within the epiblast for morphogenesis during both epiboly and neurulation and suggest that the mechanisms that drive epiboly possess common elements with those that underlie convergence and extension. (+info)
Effects of substance P in the amygdala, ventromedial hypothalamus, and periaqueductal gray on fear-potentiated startle.
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Interaction of physalaemin, substance P, and eledoisin with specific membrane receptors on pancreatic acinar cells.
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We have prepared 125I-labeled physalaemin and have examined the kinetics, stoichiometry, and chemical specificity with which the labeled peptide binds to dispersed acini from guinea pig pancreas. Binding of 125I-labeled physalaemin was saturable, temperature-dependent, and reversible and reflected interaction of the labeled peptide with a single class of binding sites on the plasma membrane of pancreatic acinar cells. Each acinar cell possessed approximately 500 binding sites, and binding of the tracer to these sites could be inhibited by physalaemin [concentration for half-maximal effect (Kd), 2 nM], substance P (Kd, 5 nM), or eledoisin (Kd, 300 nM) but not by cholecystokinin, caerulein, bombesin, litorin, gastrin, secretin, vasoactive intestinal peptide, glucagon, somatostatin, neurotensin, bovine pancreatic polypeptide, leucine-enkephalin, methionine-enkephalin, atropine, or carbamylcholine. With physalaemin, substance P, and eledoisin, there was a close correlation between the relative potency for inhibition of binding of labeled physalaemin and that for stimulation of amylase secretion. For a given peptide, however, a 3-fold higher concentration was required for half-maximal inhibition of binding than for half-maximal stimulation of amylase secretion, calcium outflux, or cyclic GMP accumulation. These results indicate that dispersed acini from guinea pig pancreas possess a single class of receptors that interact with physalaemin, substance P, and eledoisin and that occupation of 45% of these receptors will cause a maximal biological response. (+info)