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Query: UMLS:C0848283 (rundown)
 502 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Wide-tipped, low-resistance (approximately 1 M omega) pipettes were used to record the whole-cell Cl- current activated by cAMP-dependent protein kinase (PKA) in guinea-pig ventricular myocytes internally dialysed with or without GTP. Without GTP in the pipette, the response to 1 microM-isoprenaline declined with time and eventually disappeared, usually within approximately 20 min of rupturing the membrane and beginning cell dialysis. 2. This rundown of the isoprenaline response occurred more quickly with wider, lower-resistance pipette tips. 3. After complete rundown of the isoprenaline response, histamine (10 microM), another agonist known to elicit the Cl- current, also had no effect, but extracellular forskolin (1 microM) or intrapipette cAMP (1 mM) could still readily elicit the Cl- current. 4. In contrast, with 100 microM-GTP in the pipette, the response to 1 microM-isoprenaline was well maintained for periods greater than 20 min. But, if GTP was then withdrawn from the pipette, a rundown of the isoprenaline response was seen comparable to that in the experiments begun with GTP-free pipette solution. Moreover, in experiments begun without pipette GTP, the addition of 100 microM-GTP to the pipette solution, after the response to isoprenaline had disappeared, was able to restore that Cl- current response. 5. With GTP in the pipette, the forskolin-induced Cl- current could be suppressed by concurrent exposure to carbachol (10 microM). That inhibition was not seen in myocytes pretreated with pertussis toxin. In untreated myocytes dialysed with GTP-free pipette solution, after disappearance of the isoprenaline response, the muscarinic receptor-mediated inhibition was itself abolished. 6. We confirm that both beta-adrenoceptor-mediated activation of the Cl- current by isoprenaline, and muscarinic receptor-mediated inhibition of the forskolin-induced Cl- current, are mediated by G proteins, and conclude that the disappearance of both receptor-mediated responses during whole-cell recording with GTP-free pipette solution reflects the fall of cellular [GTP] below the level required to maintain G protein-dependent signal transduction.
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PMID:Pipette GTP is essential for receptor-mediated regulation of Cl- current in dialysed myocytes from guinea-pig ventricle. 133 50

The hypothalamic peptide somatostatin (SRIF) suppresses secretory activity in phenotypically distinct pituitary endocrine cells. We have used tight-seal whole-cell recording techniques to study the peptide's effects on the electrical properties of tumor pituitary cells derived from rat (GH3/B6) and human adenomas that secrete human PRL in a SRIF-sensitive manner. Both cell types exhibited qualitatively similar electrophysiological properties and electrical responses to SRIF. Under the experimental conditions employed the majority of cells spontaneously generated Ca2+-dependent actions potentials. The actions of the peptide on cellular excitability were markedly affected by the presence of horse and fetal calf sera. Without these additives the electrical responses faded and could not be studied in detail. Therefore, recordings were conducted in media containing sera. In the presence of sera almost all cells spontaneously generated Ca2+ action potentials, and peptide-induced changes in excitability were well preserved. SRIF depressed spontaneous and evoked action potential activity in a dose-dependent manner at concentrations that reduced intracellular free calcium ([Ca2+]i) and suppressed basal PRL release. Current and voltage clamp experiments revealed coordinate actions of the peptide on excitable membrane properties. SRIF (1 nM) enhanced a depolarization-activated, rapidly inactivating outward K+ current, thereby effectively reducing the rate at which action potentials occurred. Over the 10-1000 nM range SRIF slowly activated a virtually noninactivating K+ conductance over a wide range of membrane potential. This effectively hyperpolarized cells away from the threshold for triggering Ca2+-dependent action potentials and shunted the membrane. The peptide induced K+ conductance activated at the level of the resting potential was progressively lost during the intracellular dialysis of whole-cell recording. Dilute aqueous lysates of cells included in the patch pipette prevented much of the rundown of this SRIF-induced electrical response while inclusion of an ATP-regenerating system preserved some of the peptide action. Over the 10-100 nM concentration range SRIF also reduced voltage-dependent Ca2+ current. Furthermore, pretreatment of cells with pertussis toxin abolished SRIF action on cellular excitability, suggesting that SRIF can regulate the function of ionic channels through GTP-binding proteins (G proteins). The results demonstrate that SRIF acts coordinately on the primary conductances expressed in tumor PRL cells to attenuate or block Ca2+ action potential generation and thus Ga2+ entry from extracellular sources.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Somatostatin blocks Ca2+ action potential activity in prolactin-secreting pituitary tumor cells through coordinate actions on K+ and Ca2+ conductances. 245 3

Ca-dependent K and Cl currents were measured in isolated cells from rat lacrimal glands using the tight-seal whole-cell recording method. Upon application of acetylcholine (ACh), both K and Cl-selective currents were activated. The size of the ACh-activated currents declined after a few minutes of whole-cell recording. The rundown curve was composed of an initial stable period followed by a rather rapid decline. Both the length of the initial plateau and the speed of the falling phase were dependent on cell size and recording pipette resistance. The results suggest that the rundown was due to washout of an unknown cytosolic substance. Another manifestation of washout was an increase in the delay of the response. Plots of the inverse of the delay as a function of time in whole-cell recording showed again an initial plateau and a falling phase, but the stable period lasted less than in amplitude plots. Analysis of the washout time course suggested that the cytosolic substance has a diffusion coefficient of 5.4 x 10(-6) cm2/s, corresponding to a molecular weight of 350. Washed-out cells were insensitive to GTP-gamma-S, but responded normally to an internal application of inositol-trisphosphate (InsP3), introduced through the pipette. Thus, the step of the response which is sensitive to washout is closely related to the production of InsP3. Addition of various exogenous water soluble substances failed to halt washout. Among the inactive substances were GTP (or a combination of Mg and GTP) and small water soluble precursors of InsP3. The results imply that the production of InsP3 by muscarinic agonists in exocrine glands requires the presence of a small molecular weight, water soluble substance. It is suggested that this substance is an unknown co-factor of phospholipase C or of Gp, the GTP binding protein governing the production of InsP3.
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PMID:Diffusion into the patch-clamp recording pipette of a factor necessary for muscarinic current response. 270 3

The Ca2+ current recorded by the whole-cell technique in chromaffin cells shows, before the often described rundown, a transient facilitation or runup. Initial current amplitude was 570 +/- 165 pA and then it increased by 49 +/- 23% (n = 19, SD) over 2 +/- 1 min in the absence of adenosine 5'-triphosphate (ATP). In the presence of ATP, this process occurred with the same magnitude but it was slowed in a dose-dependent manner, lasting 17 +/- 2 min with 2 mM ATP (n = 8). Since adenosine 5'-diphosphate (ADP) does not reproduce this ATP effect, a complex series of phosphorylations is likely to intervene and we show that, at least, a cAMP-dependent i.e., cyclic adenosine monophosphate) phosphorylation occurs. Pertussis toxin (PTX) pretreatment yielded an already maximal Ca2+ current (around 1000 pA) at the time of the patch rupture, which only slightly increased thereafter (10%, n = 11). Also, guanosine 5'-diphosphate (GDP) and guanosine 5'-O-(2-thiodiphosphate) (GDP[ beta s]), induced a fast runup, which was absent in the presence of GTP. Furthermore, we show that facilitation does not occur in the presence of dihydrophyridine (DHP) antagonists. Globally, our data suggest that an ATP-dependent phosphorylation stabilizes the inhibitory control exerted by a PTX-sensitive G protein and, as a result, slows down the facilitation of L-type Ca2+ channels. The recruitment of L-type channels can also be facilitated by the application of a DHP agonist or a depolarizing prepulse protocol.l We show that these processes are only effective over a period which parallels the runup and are not additive to it.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:ATP and G proteins affect the runup of the Ca2+ current in bovine chromaffin cells. 749 Dec 66

The basophilic leucaemia cell line RBL-2H3 exhibits a robust inwardly rectifying potassium current, IKIR, which is likely to be modulated by G proteins. We examined the physiological and molecular properties of this KIR conductance to define the nature of the underlying channel species. The macroscopic conductance revealed characteristics typical of classical K+ inward rectifiers of the IRK type. Channel gating was rapid, first order (tau approximately 1 ms at -100 mV) and steeply voltage dependent. Both activation potential and slope conductance were dependent on extracellular K+ concentration ([K+]o) and inward rectification persisted in the absence of internal Mg2+. The current was susceptible to a concentration- and voltage-dependent block by extracellular Na+, Cs+ and Ba2+. Initial IKIR whole-cell amplitudes as well as current rundown were dependent on the presence of 1 mM internal ATP. Perfusion of intracellular guanosine 5'-Q-(3-thiotriphosphate) (GTP[gamma S]) suppressed IKIR with an average half-time of decline of approximately 400 s. It was demonstrated that the dominant IRK-type 25 pS conductance channel was indeed suppressed by 100 microM preloaded GTP[gamma S]. Reverse transcriptase-polymerase chain reactions (RT-PCR) with RBL cell poly(A)+ RNA identified a full length K+ inward rectifier with 94% base pair homology to the recently cloned mouse IRK1 channel. It is concluded that RBL cells express a classical voltage-dependent IRK-type K+ inward rectifier RBL-IRK1 which is negatively controlled by G proteins.
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PMID:Physiological and molecular characterization of an IRK-type inward rectifier K+ channel in a tumour mast cell line. 760 35

1. The effect of G protein activation on the ATP-sensitive K+ (K+ATP) channel was examined in inside-out patches from guinea-pig ventricular myocytes. At low (0.3 mM) intracellular ATP concentration ([ATP]i) in the bathing solution, in the absence of agonists in the pipette, guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) or AlF4- applied to the intracellular side of the patch membrane gradually activated the K+ATP channel. The activation by GTP gamma S was irreversible, although high [ATP]i could completely close the channel. 2. In ATP-free media GTP gamma S did not increase further the activity of the fully active channel, and was unable to reactivate the channel in the non-operative state after rundown. [ATP]i-channel activity curves constructed before and after GTP gamma S application demonstrated that GTP gamma S shifts the half-inhibitory [ATP]i from 19.5 to 110 microM without changing the Hill coefficient. 3. When acetylcholine or adenosine was included in the pipette, intracellular GTP reversibly activated the K+ATP channel which was partially inhibited by [ATP]i. 4. These results indicate that G protein may stimulate myocardial K+ATP channels in the operative state by reducing the potency of ATP inhibition. The possible coupling of the G protein with muscarinic as well as A1 adenosine receptors is suggested.
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PMID:Mode of regulation by G protein of the ATP-sensitive K+ channel in guinea-pig ventricular cell membrane. 796 25

We have investigated the coupling between opioid receptors and different types of Ca2+ channels in neurons acutely isolated from the nucleus tractus solitarius (NTS) of the rat. Using fura-2-based imaging we found that Ca2+ transients evoked by depolarization with 50 mM KCl were suppressed by the mu-opioid receptor agonist D-Ala2,N-MePhe4,Gly5-ol-enkephalin (DAMGO) and less effectively by the kappa-receptor agonist U-69,593. The delta-receptor agonist D-Pen2,D-Pen5-enkephalin (DPDPE) was ineffective. In whole-cell voltage-clamp recordings from these neurons, depolarizing voltage steps elicited high-threshold Ca2+ currents that could be distinguished pharmacologically into different components. Part of the current could be blocked by dihydropyridines, part by omega-conotoxin-GVIA and part by omega-agatoxin-IVA. This suggests that the neurons contained L-, N-, and P/Q-type Ca2+ channels. DAMGO and U-69,593 both blocked part of the Ca2+ current but DPDPE was ineffective. Perfusion of GTP-gamma-S into the cells produced a rapid rundown of the Ca2+ current and occluded further effects of the opioid agonists, suggesting the involvement of a G-protein in the coupling mechanism. Inhibition of L-channels did not alter the effect of DAMGO. On the other hand inhibition of N-channels occluded about 80% of the effect of DAMGO. Inhibition of the P/Q-current occluded the remainder of the DAMGO effect. Thus, it appears that activation of opioid receptors can inhibit N- and P/Q-type Ca2+ channels but not L-channels in these cells. It is likely that such effects are important in opioid-mediated inhibition of transmitter release in the brain.
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PMID:Opioid receptors modulate diverse types of calcium channels in the nucleus tractus solitarius of the rat. 799 99

We examined the effect of altering the cytoskeleton polymerization state by treatment with nocodazole and taxol on glycine-evoked currents in patch-clamp recordings from cultured spinal cord neurones. Adding ATP and GTP to the pipette solution did not prevent the rundown of the peak current. In the absence or in the presence of ATP, the proportion of the non-desensitizing part of the glycine evoked-current declined with time. Adding intracellular GTP and ATP stabilized glycine-evoked responses although the proportion of non-inactivating current was reduced. Nocodazole reduced by itself the proportion of the non-inactivating current whereas taxol (with ATP and GTP) had an opposite effect. These results suggest that the polymerization state of microtubules has functional consequences on glycine receptors.
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PMID:Effects of nocodazole and taxol on glycine evoked currents on rat spinal cord neurones in culture. 854 1

Mast cells permeabilized by treatment with streptolysin-O in the presence of Ca2+ and GTP-gamma-S can secrete almost 100% of their contained N-acetyl-beta-D-glucosaminidase. If these stimuli are provided to the permeabilized cells after a delay, the response is diminished and the ability of the cells to undergo secretion runs down progressively over a period of about 30 min. This is thought to be due to the loss of key proteins involved in the exocytotic mechanism. Using this effect as the basis of a biological assay, we have isolated a protein from bovine brain cytosol that retards the loss of responsiveness to stimulation by Ca2+ and GTP-gamma-S. Purification of this protein and peptide sequencing have enabled us to identify it as the small GTP-binding protein rac complexed to the guanine nucleotide exchange inhibitor rhoGDI. Both proteins are required to retard the loss of the secretory response, while purified rhoGDI applied alone accelerates the rundown.
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PMID:Purification and identification of FOAD-II, a cytosolic protein that regulates secretion in streptolysin-O permeabilized mast cells, as a rac/rhoGDI complex. 886 68

We investigated the action of adenosine and GTP on KATP channels, using inside-out patch clamp recordings from dissociated single fibers of rat flexor digitorum brevis (FDB) skeletal muscle. In excised patches, KATP channels could be activated by a combination of an extracellular adenosine agonist and intracellular Mg2+-ATP and GTP or GTP-gamma-S. The activation required hydrolyzable ATP and could be partially reversed with Mg2+, suggesting that it may involve a G-protein dependent phosphorylation of KATP channels. We found that KATP channels of the rat FDB could not be activated by Mg2+-ATP alone or by Mg2+-ATP in the presence of extracellular adenosine. Patches whose channel activity had been 'rundown' by Ca2+ could not be recovered by adenosine, GTP or Mg2+-ATP. KATP channels activated by adenosine receptor agonists had a similar ATP sensitivity to those under control conditions; but adenosine appears to be able to switch these KATP channels from an inactive to an active mode.
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PMID:Effect of adenosine and intracellular GTP on KATP channels of mammalian skeletal muscle. 913 22


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