RESUMEN
BACKGROUND: Biological clocks time many physiological parameters with periodicities close to 24 h; those which persist in the absence of environmental cues are circadian. An earlier shuttle experiment (STS-9) examined circadian pacemaker function and growth rate of Neurospora crassa and demonstrated damped rhythm amplitudes, increased variability in period lengths and altered growth rates. HYPOTHESIS: Postflight studies suggested that accelerative forces of launch could have induced rhythm alterations. Differences in growth rate may have been due to an alteration of metabolic rate. METHODS: Race tubes inoculated with bd or csp strains were flown aboard STS-32, exposed to ambient mid-deck temperatures throughout flight, and exposed to light only during marking procedures. Period, rhythm amplitude, and growth rate were determined and compared to orbital environmental controls (OES) and 25 degrees C ground controls (GC). RESULTS: Unlike the previous flight exposurement, the rhythm persisted quite normally. bd flight and OES cultures each displayed lengthened periods of a similar magnitude when compared to GC. The lengthened periods of csp flight cultures while longer than GC, were shorter than OES. Shuttle temperatures were relatively warm, however the increased period length in space was greater than predicted by the known Q10. Growth rates also increased substantially during flight, which could not be accounted for by thermal mechanisms alone. CONCLUSION: It is likely that some of the cultures may have entrained to the unexpected diurnal temperature variations; however, other cultures did not entrain, yet retained rhythmicity with increased periods. The results also suggest an increased metabolic rate during spaceflight.
Asunto(s)
Ritmo Circadiano , Neurospora crassa/crecimiento & desarrollo , Vuelo EspacialRESUMEN
This study sought to characterize the action of neurokinin B (NKB) and senktide, a selective synthetic agonist for NK3 receptors, on the myenteric plexus of the guinea pig small intestine. Both peptides stimulated a dose-dependent release of [3H]-acetylcholine (ACh). The mean effective dose values were 1 x 10(-9) for NKB and 3 x 10(-11) M for senktide. The action of these two neurokinins was blocked by the removal of Ca2+ and was sensitive to tetrodotoxin. The release of [3H]ACh was antagonized by omega-conotoxin GVIA, implying the involvement of N-type voltage-sensitive calcium channels. Senktide-evoked ACh release was also insensitive to nifedipine or flunarizine but was blocked by diltiazem. Treatment with protein kinase C (PKC) inhibitors (H-7 and polymyxin B) or activators (12-tetradecanoylphorbol 13-acetate and SC-9) failed to alter the NK3 receptor-mediated ACh output. Our data did not support an action mediated via PKC upon the activation of NK3 receptors on myenteric cholinergic neurons.
Asunto(s)
Acetilcolina/metabolismo , Plexo Mientérico/metabolismo , Receptores de Neurotransmisores/fisiología , Animales , Calcio/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Activación Enzimática , Cobayas , Técnicas In Vitro , Neuroquinina B/farmacología , Fragmentos de Péptidos/farmacología , Proteína Quinasa C/antagonistas & inhibidores , Proteína Quinasa C/metabolismo , Receptores de Neuroquinina-3 , Receptores de Neurotransmisores/metabolismo , Sustancia P/análogos & derivados , Sustancia P/farmacología , Tetrodotoxina/farmacologíaRESUMEN
Release of [3H]acetylcholine ([3H]ACh) was examined in a submucous plexus preparation obtained from the guinea pig small intestine in vitro. Constant-current field stimulation evoked ACh output; this output was dependent on the stimulus frequency applied. Maximal release was observed at 10 Hz; this release was blocked by tetrodotoxin (1 x 10(-6) M) or in Ca2(+)-free buffer. Serotonin [5-hydroxytryptamine (5-HT)] stimulated the release of ACh dose dependently, with an ED50 of 5 x 10(-7) M. Substance P was ineffective, while vasoactive intestinal peptide weakly stimulated ACh secretion. Several neuropeptides were tested on their ability to modulate 5-HT-evoked ACh release. Dynorphin A inhibited 5-HT-stimulated ACh release, while Met-enkephalin was without any effect. Both somatostatin and galanin were effective modulators, with an inhibitory effect in the submicromolar range and an excitatory effect at higher concentrations. The response characteristics of the cholinergic neurons of submucosal plexus differ markedly from those of the myenteric plexus. These distinct features form an important framework for future functional studies on submucous plexus neurons.
Asunto(s)
Acetilcolina/fisiología , Íleon/inervación , Yeyuno/inervación , Plexo Mientérico/fisiología , Neuronas/fisiología , Neuropéptidos/farmacología , Serotonina/farmacología , Animales , Dinorfinas/farmacología , Estimulación Eléctrica , Encefalina Metionina/farmacología , Femenino , Galanina , Técnicas In Vitro , Músculo Liso/inervación , Neuronas/efectos de los fármacos , Péptidos/farmacología , Conejos , Somatostatina/farmacología , Sustancia P/farmacología , Péptido Intestinal Vasoactivo/farmacologíaRESUMEN
Longitudinal muscle strips adhered with myenteric plexus were subjected to enzyme digestion under controlled conditions in a Krebs-bicarbonate buffer solution containing a mixture of collagenase, deoxyribonuclease, protease, choline chloride, and bovine serum albumin for 30 min at 37 degrees C. Myenteric ganglia, singly or in multiple aggregates, were harvested with micropipette and labeled with [3H]choline for [3H]acetylcholine (ACh) release studies. When examined by light or electron (transmission or scanning) microscopy, the ganglia exhibited their normal structural characteristics with axon bundles, dendrites, cell bodies, and vesiculated processes. Depolarization with elevated potassium or veratrine hydrochloride significantly elevated the efflux of [3H] ACh. Perfusion with tachykinins (substance P and substance K), vasoactive intestinal peptide, forskolin, or serotonin also significantly increased the release of [3H]ACh. This study demonstrated that enzyme-dissociated myenteric ganglia, notably free of muscle or connective tissue components, were structurally well preserved and were amenable to functional studies targeted specifically for the enteric plexus neurons.
Asunto(s)
Acetilcolina/metabolismo , Desoxirribonucleasa I/metabolismo , Colagenasa Microbiana/metabolismo , Plexo Mientérico/metabolismo , Péptido Hidrolasas/metabolismo , 1-(5-Isoquinolinesulfonil)-2-Metilpiperazina , Animales , Calcio/farmacología , Colforsina/farmacología , Femenino , Cobayas , Isoquinolinas/farmacología , L-Lactato Deshidrogenasa/metabolismo , Microscopía Electrónica , Microscopía Electrónica de Rastreo , Plexo Mientérico/efectos de los fármacos , Plexo Mientérico/ultraestructura , Neuropéptidos/farmacología , Piperazinas/farmacología , Potasio/farmacología , Proteína Quinasa C/antagonistas & inhibidores , Serotonina/farmacología , Tetrodotoxina/farmacología , Veratrina/farmacologíaRESUMEN
Forskolin, an activator of adenylate cyclase, was used to examine the regulation of [3H]acetylcholine (ACh) release by cyclic AMP (cAMP)-related mechanisms in myenteric plexus-longitudinal muscle preparations of guinea pig small intestine. Forskolin evoked a dose-related increase in [3H]ACh release. Both dibutyryl-cAMP and 8-Br-cAMP significantly elevated [3H]ACh secretion. In the presence of phosphodiesterase inhibitors (theophylline and 3-isobutyl-1-methylxanthine), the basal [3H]ACh output was increased. There was a significantly greater stimulation when forskolin was used to incite endogenous cAMP synthesis and phosphodiesterase inhibitors were simultaneously applied to prevent cAMP breakdown. The enhancement of forskolin-stimulated release by theophylline or 3-isobutyl-1-methylxanthine strongly implicates a synergistic interaction between the two. These findings suggest that forskolin acts to increase ACh release by a modulation of endogenous cAMP and further support a cAMP-mediated mechanism in the secretion of ACh from myenteric cholinergic neurons.
Asunto(s)
Acetilcolina/metabolismo , Colforsina/farmacología , AMP Cíclico/farmacología , Plexo Mientérico/efectos de los fármacos , 1-Metil-3-Isobutilxantina/farmacología , 8-Bromo Monofosfato de Adenosina Cíclica/farmacología , Animales , Bucladesina/farmacología , Cobayas , Técnicas In Vitro , Intestino Delgado/inervación , Contracción Muscular/efectos de los fármacos , Plexo Mientérico/metabolismo , Teofilina/farmacologíaRESUMEN
The effect of galanin on the [3H]ACh release from myenteric plexus-longitudinal muscle strips of the guinea pig small intestine was studied. While galanin did not alter the basal spontaneous efflux of ACh, it significantly depressed the ACh release evoked by electrical stimulation or caused by VIP and substance P. These results suggest an important neuromodulatory role for galanin in the enteric nervous system.
Asunto(s)
Acetilcolina/metabolismo , Fibras Colinérgicas/efectos de los fármacos , Inhibidores de la Colinesterasa , Intestino Delgado/inervación , Plexo Mientérico/efectos de los fármacos , Péptidos/farmacología , Animales , Fibras Colinérgicas/fisiología , Estimulación Eléctrica , Galanina , Cobayas , Técnicas In Vitro , Intestino Delgado/efectos de los fármacos , Plexo Mientérico/fisiología , Sustancia P/farmacología , Péptido Intestinal Vasoactivo/farmacologíaRESUMEN
Release of [3H]acetylcholine (ACh) under the influence of bradykinin was measured from myenteric plexus-longitudinal muscle strips taken from guinea pig small intestine. Bradykinin stimulated the efflux of [3H]ACh in a dose-dependent manner. This stimulation by bradykinin was resistant to the effect of [Des-Arg9-Leu8]-bradykinin but not to indomethacin, indicating that the ACh-releasing action of bradykinin was mediated indirectly by a prostaglandin mechanism. Direct evidence for a stimulation of ACh release by prostaglandin E1 was obtained. Prostaglandin was able to stimulate ACh release in a dose-related fashion. The inhibition of bradykinin-induced ACh release by indomethacin was partly reversed by exogenous prostaglandin E1. These results suggest a neuromodulatory role for bradykinin in the enteric nervous system.
Asunto(s)
Acetilcolina/metabolismo , Alprostadil/farmacología , Bradiquinina/farmacología , Plexo Mientérico/metabolismo , Animales , Bradiquinina/análogos & derivados , Cobayas , Técnicas In Vitro , Indometacina/farmacología , Intestino Delgado/inervación , Cinética , Plexo Mientérico/efectos de los fármacos , Prostaglandinas/fisiologíaRESUMEN
This study reports on a difference in the inhibitory action of the neuropeptides somatostatin and Met-enkephalin on acetylcholine (ACh) release from myenteric plexus-longitudinal muscle strips of guinea pig small intestine. Met-enkephalin (8.7 X 10(-8) M) inhibited ACh release evoked by either substance P (3.7 X 10(-8) M) or neurotensin (7.5 X 10(-11) M), and this inhibition could be reversed by naloxone (5 X 10(-8) to 5 X 10(-5) M). Neurotensin-induced ACh release was also sensitive to the inhibition by somatostatin. However, when tested in a dose range from 6.1 X 10(-8) to 6.1 X 10(-6) M, somatostatin was ineffective in reducing the efflux of ACh evoked by substance P. These observations provided evidence to support the view that inhibitory peptidergic neurons within the myenteric plexus modulate the activity of cholinergic neurons with a high degree of specificity and that both somatostatin and Met-enkephalin have distinct neuromodulatory functions in the gut.
Asunto(s)
Plexo Mientérico/efectos de los fármacos , Somatostatina/farmacología , Acetilcolina/metabolismo , Animales , Encefalina Metionina/farmacología , Femenino , Cobayas , Intestino Delgado/efectos de los fármacos , Intestino Delgado/metabolismo , Plexo Mientérico/metabolismo , Naloxona/farmacología , Neurotensina/farmacología , Sustancia P/farmacologíaRESUMEN
In the guinea-pig myenteric plexus-longitudinal muscle preparation from the small intestine, the release of acetylcholine evoked by substance P and vasoactive intestinal polypeptide was examined in vitro. Both neuropeptides stimulated efflux of [3H]acetylcholine from myenteric neurons in a calcium-dependent manner. This observation is consistent with the view that neurotransmitter release elicited by neuropeptides requires the presence of extracellular calcium ions.