首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
  1. Relaxation of the methoxamine-precontracted rat small mesenteric artery by endothelium-derived hyperpolarizing factor (EDHF) was compared with relaxation to the cannabinoid, anandamide (arachidonylethanolamide). EDHF was produced in a concentration- and endothelium-dependent fashion in the presence of NG-nitro-L-arginine methyl ester (L-NAME, 100 μM) by either carbachol (pEC50 [negative logarithm of the EC50]=6.19±0.01, Rmax [maximum response]=93.2±0.4%; n=14) or calcium ionophore A23187 (pEC50=6.46±0.02, Rmax=83.6±3.6%; n=8). Anandamide responses were independent of the presence of endothelium or L-NAME (control with endothelium: pEC50=6.31±0.06, Rmax=94.7±4.6%; n=10; with L-NAME: pEC50=6.33±0.04, Rmax=93.4±6.0%; n=4).
  2. The selective cannabinoid receptor antagonist, SR 141716A (1 μM) caused rightward shifts of the concentration-response curves to both carbachol (2.5 fold) and A23187 (3.3 fold). It also antagonized anandamide relaxations in the presence or absence of endothelium giving a 2 fold shift in each case. SR 141716A (10 μM) greatly reduced the Rmax values for EDHF-mediated relaxations to carbachol (control, 93.2±0.4%; SR 141716A, 10.7±2.5%; n=5; P<0.001) and A23187 (control, 84.8±2.1%; SR 141716A, 3.5±2.3%; n=6; P<0.001) but caused a 10 fold parallel shift in the concentration-relaxation curve for anandamide without affecting Rmax.
  3. Precontraction with 60 mM KCl significantly reduced (P<0.01; n=4 for all) relaxations to 1 μM carbachol (control 68.8±5.6% versus 17.8±7.1%), A23187 (control 71.4±6.1% versus 3.9±0.45%) and anandamide (control 71.1±7.0% versus 5.2±3.6%). Similar effects were seen in the presence of 25 mM K+. Incubation of vessels with pertussis toxin (PTX; 400 ng ml−1, 2 h) also reduced (P<0.01; n=4 for all) relaxations to 1 μM carbachol (control 63.5±7.5% versus 9.0±3.2%), A23187 (control 77.0±5.8% versus 16.2±7.1%) and anandamide (control 89.8±2.2% versus 17.6±8.7%).
  4. Incubation of vessels with the protease inhibitor phenylmethylsulphonyl fluoride (PMSF; 200 μM) significantly potentiated (P<0.01), to a similar extent (∼2 fold), relaxation to A23187 (pEC50: control, 6.45±0.04; PMSF, 6.74±0.10; n=4) and anandamide (pEC50: control, 6.31±0.02; PMSF, 6.61±0.08; n=8). PMSF also potentiated carbachol responses both in the presence (pEC50: control, 6.25±0.01; PMSF, 7.00±0.01; n=4; P<0.01) and absence (pEC50: control, 6.41±0.04; PMSF, 6.88±0.04; n=4; P<0.001) of L-NAME. Responses to the nitric oxide donor S-nitroso-N-acetylpenicillamine (SNAP) were also potentiated by PMSF (pEC50: control, 7.51±0.06; PMSF, 8.00±0.05, n=4, P<0.001).
  5. EDHF-mediated relaxation to carbachol was significantly attenuated by the K+ channel blocker tetraethylammonium (TEA; 1 mM) (pEC50: control, 6.19±0.01; TEA, 5.61±0.01; n=6; P<0.01). In contrast, TEA (1 mM) had no effect on EDHF-mediated relaxation to A23187 (pEC50: control, 6.47±0.04; TEA, 6.41±0.02, n=4) or on anandamide (pEC50: control, 6.28±0.06; TEA, 6.09±0.02; n=5). TEA (10 mM) significantly (P<0.01) reduced the Rmax for anandamide (control, 94.3±4.0%; 10 mM TEA, 60.7±4.4%; n=5) but had no effect on the Rmax to carbachol or A23187.
  6. BaCl2 (100 μM), considered to be selective for blockade of inward rectifier K+ channels, had no significant effect on relaxations to carbachol or A23187, but caused a small shift in the anandamide concentration-response curve (pEC50: control, 6.39±0.01; Ba2+, 6.20±0.01; n=4; P<0.01). BaCl2 (1 mM; which causes non-selective block of K+ channels) significantly (P<0.01) attenuated relaxations to all three agents (pEC50 values: carbachol, 5.65±0.02; A23187, 5.84±0.04; anandamide, 5.95±0.02; n=4 for each).
  7. Apamin (1 μM), a selective blocker of small conductance, Ca2+-activated, K+ channels (SKCa), 4-aminopyridine (1 mM), a blocker of delayed rectifier, voltage-dependent, K+ channels (Kv), and ciclazindol (10 μM), an inhibitor of Kv and adenosine 5′-triphosphate (ATP)-sensitive K+ channels (KATP), significantly reduced EDHF-mediated relaxations to carbachol, but had no significant effects on A23187 or anandamide responses.
  8. Glibenclamide (10 μM), a KATP inhibitor and charybdotoxin (100 or 300 nM), a blocker of several K+ channel subtypes, had no significant effect on relaxations to any of the agents. Iberiotoxin (50 nM), an inhibitor of large conductance, Ca2+-activated, K+ channels (BKCa), had no significant effect on the relaxation responses, either alone or in combination with apamin (1 μM). Also, a combination of apamin (1 μM) with either glibenclamide (10 μM) or 4-aminopyridine (1 mM) did not inhibit relaxation to carbachol significantly more than apamin alone. Neither combination had any significant effect on relaxation to A23187 or anandamide.
  9. A combination of apamin (1 μM) with charybdotoxin (100 nM) abolished EDHF-mediated relaxation to carbachol, but had no significant effect on that to A23187. Apamin (1 μM) and charybdotoxin (300 nM) together consistently inhibited the response to A23187, while apamin (1 μM) and ciclazindol (10 μM) together inhibited relaxations to both carbachol and A23187. None of these toxin combinations had any significant effect on relaxation to anandamide.
  10. It was concluded that the differential sensitivity to K+ channel blockers of EDHF-mediated responses to carbachol and A23187 might be due to actions on endothelial generation of EDHF, as well as its actions on the vascular smooth muscle, and suggests care must be taken in choosing the means of generating EDHF when making comparative studies. Also, the relaxations to EDHF and anandamide may involve activation of cannabinoid receptors, coupled via PTX-sensitive G-proteins to activation of K+ conductances. The results support the hypothesis that EDHF is an endocannabinoid but relaxations to EDHF and anandamide show differential sensitivity to K+ channel blockers, therefore it is likely that anandamide is not identical to EDHF in the small rat mesenteric artery.
  相似文献   

2.
  1. The effects of nifedipine on both levcromakalim-induced membrane currents and unitary currents in pig proximal urethra were investigated by use of patch-clamp techniques (conventional whole-cell configuration and cell-attached patches).
  2. Nifedipine had a voltage-dependent inhibitory effect on voltage-dependent Ba2+ currents at −50 mV (Ki=30.6 nM).
  3. In current-clamp mode, subsequent application of higher concentrations of nifedipine (⩾30 μM) caused a significant depolarization even after the membrane potential had been hyperpolarized to approximately −82 mV by application of 100 μM levcromakalim.
  4. The 100 μM levcromakalim-induced inward current (symmetrical 140 mM K+ conditions, −50 mV) was inhibited by additional application of three different types of Ca antagonists (nifedipine, verapamil and diltiazem, all at 100 μM). In contrast, Bay K 8644 (1 μM) possessed no activating effect on the amplitude of this glibenclamide-sensitive current.
  5. When 100 μM nifedipine was included in the pipette solution during conventional whole-cell recording at −50 mV, application of levcromakalim (100 μM) caused a significant inward membrane current which was suppressed by 5 μM glibenclamide. On the other hand, inclusion of 5 μM glibenclamide in the pipette solution prevented levcromakalim from inducing an inward membrane current.
  6. The levcromakalim-induced K+ channel openings in cell-attached configuration were suppressed by subsequent application of 5 μM glibenclamide but not of 100 μM nifedipine.
  7. These results suggest that in pig proximal urethra, nifedipine inhibits the glibenclamide-sensitive 43 pS K+ channel activity mainly through extracellular blocking actions on the K+ channel itself.
  相似文献   

3.
  1. To determine the contribution of the various phosphodiesterase (PDE) isozymes to the regulation of the L-type calcium current (ICa(L)) in the human myocardium, we investigated the effect of selective and non-selective PDE inhibitors on ICa(L) in single human atrial cells by use of the whole-cell patch-clamp method. We repeated some experiments in rabbit atrial myocytes, to make a species comparison.
  2. In human atrial cells, 100 μM pimobendan increased ICa(L) (evoked by depolarization to +10 mV from a holding potential of −40 mV) by 250.4±45.0% (n=15), with the concentration for half-maximal stimulation (EC50) being 1.13 μM. ICa(L) was increased by 100 μM UD-CG 212 by 174.5±30.2% (n=10) with an EC50 value of 1.78 μM in human atrial cells. These two agents inhibit PDE III selectively.
  3. A selective PDE IV inhibitor, rolipram (1–100 μM), did not itself affect ICa(L) in human atrial cells. However, 100 μM rolipram significantly enhanced the effect of 100 μM UD-CG 212 on ICa(L) (increase with UD-CG 212 alone, 167.9±33.9, n=5; increase with the two agents together, 270.0±52.2%; n=5, P<0.05). Rolipram also enhanced isoprenaline (5 nM)-stimulated ICa(L) by 52.9±9.3% (n=5) in human atrial cells.
  4. In rabbit atrial cells, ICa(L) at +10 mV was increased by 22.1±9.0% by UD-CG 212 (n=10) and by 67.4±12.0% (n=10) by pimobendan (each at 100 μM). These values were significantly lower than those obtained in human atrial cells (P<0.0001). Rolipram (1–100 μM) did not itself affect ICa(L) in rabbit atrial cells. However, ICa(L) was increased by 215.7±65.2% (n=10) by the combination of 100 μM UD-CG 212 and 100 μM rolipram. This value was almost 10 times larger than that obtained for the effect of 100 μM UD-CG 212 alone.
  5. These results imply a species difference: in the human atrium, the PDE III isoform seems dominant, whereas PDE IV may be more important in the rabbit atrium for regulating ICa(L). However, PDE IV might contribute significantly to the regulation of intracellular cyclic AMP in human myocardium when PDE III is already inhibited or when the myocardium is under β-adrenoceptor-mediated stimulation.
  相似文献   

4.
  1. The rat μ-opioid receptor has recently been cloned, yet its second messenger coupling remains unclear. The endogenous μ-opioid receptor in SH-SY5Y cells couples to phospholipase C (PLC), increases [Ca2+]i and inhibits adenylyl cyclase (AC). We have examined the effects of μ-opioid agonists on inositol(1,4,5)trisphosphate (Ins(1,4,5)P3), [Ca2+]i and adenosine 3′ : 5′-cyclic monophosphate (cyclic AMP) formation in Chinese hamster ovarian (CHO) cells transfected with the cloned μ-opioid receptor.
  2. Opioid receptor binding was assessed with [3H]-diprenorphine ([3H]-DPN) as a radiolabel. Ins(1,4,5)P3 and cyclic AMP were measured by specific radioreceptor assays. [Ca2+]i was measured fluorimetrically with Fura-2.
  3. Scatchard analysis of [3H]-DPN binding revealed that the Bmax varied between passages. Fentanyl (10 pM–1 μM) dose-dependently displaced [3H]-DPN, yielding a curve which had a Hill slope of less than unity (0.6±0.1), and was best fit to a two site model, with pKi values (% of sites) of 9.97±0.4 (27±4.8%) and 7.68±0.07 (73±4.8%). In the presence of GppNHp (100 μM) and Na+ (100 mM), the curve was shifted to the right and became steeper (Hill slope=0.9±0.1) with a pKi value of 6.76±0.04.
  4. Fentanyl (0.1 nM–1 μM) had no effect on basal, but dose-dependently inhibited forskolin (1 μM)-stimulated, cyclic AMP formation (pIC50=7.42±0.23), in a pertussis toxin (PTX; 100 ng ml−1 for 24 h)-sensitive and naloxone-reversible manner (Ki=1.7 nM). Morphine (1 μM) and [D-Ala2, MePhe4, gly(ol)5]-enkephalin (DAMGO, 1 μM) also inhibited forskolin (1 μM)-stimulated cyclic AMP formation, whilst [D-Pen2, D-Pen5], enkephalin (DPDPE, 1 μM) did not.
  5. Fentanyl (0.1 nM–10 μM) caused a naloxone (1 μM)-reversible, dose-dependent stimulation of Ins(1,4,5)P3 formation, with a pEC50 of 7.95±0.15 (n=5). PTX (100 ng ml−1 for 24 h) abolished, whilst Ni2+ (2.5 mM) inhibited (by 52%), the fentanyl-induced Ins(1,4,5)P3 response. Morphine (1 μM) and DAMGO (1 μM), but not DPDPE (1 μM), also stimulated Ins(1,4,5)P3 formation. Fentanyl (1 μM) also caused an increase in [Ca2+]i (80±16.4 nM, n=6), reaching a maximum at 26.8±2.5 s. The increase in [Ca2+]i remained elevated until sampling ended (200 s) and was essentially abolished by the addition of naloxone (1 μM). Pre-incubation with naloxone (1 μM, 3 min) completely abolished fentanyl-induced increases in [Ca2+]i.
  6. In conclusion, the cloned μ-opioid receptor when expressed in CHO cells stimulates PLC and inhibits AC, both effects being mediated by a PTX-sensitive G-protein. In addition, the receptor couples to an increase in [Ca2+]i. These findings are consistent with the previously described effector-second messenger coupling of the endogenous μ-opioid receptor.
  相似文献   

5.
  1. Interations were investigated between loreclezole, chlormethiazole and pentobarbitone as potentiators of depolarization responses mediated by γ-aminobutyric acidA (GABAA) receptors on afferent nerve terminals in the rat cuneate nucleus in vitro. These drugs were also compared as modulators of [3H]-flunitrazepam (FNZ) binding to synaptic membranes prepared from rat whole brain homogenate.
  2. In rat cuneate nucleus slices, the drugs shifted muscimol log dose–response lines to the left in an approximately parallel fashion with the result that 200 μM chlormethiazole potentiated muscimol responses by 0.567±0.037 log unit (mean±s.e.mean, n=4) while loreclezole gave a maximal potentiation at 10 μM of only 0.121±0.037 (n=6) log unit and 0.071±0.039 (n=22) at 50 μM.
  3. While 50 μM chlormethiazole and 30 μM pentobarbitone showed no significant interactions between each other when potentiating muscimol responses in combination, 50 μM loreclezole in combination with either chlormethiazole or pentobarbitone attenuated their potentiating effects, possibly by inducing desensitization of GABAA receptors.
  4. In the [3H]-FNZ binding studies on well-washed membranes, loreclezole enhanced binding to a maximum of 47.3±2.83% of control (mean±s.e.mean, n=3) at 300 μM. Scatchard analysis revealed no change in Bmax but a decrease in KD for [3H]-FNZ from 3.9±0.29 nM to 2.7±0.10 nM (mean±s.e.mean, n=4) in the presence of 100 μM loreclezole. In contrast, 100 μM chlormethiazole caused no potentiation. A small component of the enhancement by loreclezole could be blocked by 100 μM bicuculline and could also be blocked by 100 μM chlormethiazole. It seems likely that the effects on [3H]-FNZ binding are due predominantly to direct actions of the drugs on the GABAA receptor and are separate from the GABA-potentiating effects.
  5. The results indicate distinctly different profiles of action for loreclezole, chlormethiazole and pentobarbitone on GABAA receptors.
  相似文献   

6.
  1. Although extracellular adenosine 5′-triphosphate (ATP) is the natural ligand for the P2Z receptor of human lymphocytes it is less potent than 3′-O-(4-benzoylbenzoyl)-ATP (BzATP) in opening the associated ion channel, which conducts a range of permeants including Ba2+ and ethidium+. We have quantified the influx of ethidium+ into lymphocytes produced by BzATP, ATP, 2-methylthio-ATP (2MeSATP) and ATPγS, studied competition between ATP and BzATP and investigated the effects of KN-62, a new and potent inhibitor of the P2Z receptor.
  2. BzATP and ATP stimulated ethidium+ influx with EC50 values of 15.4±1.4 μM (n=5) and 85.6±8.8 μM (n=5), respectively. The maximal response to ATP was only 69.8±1.9% of that for BzATP. Hill analysis gave nH of 3.17±0.24 (n=3) and 2.09±0.45 (n=4) for BzATP and ATP, suggesting greater positive cooperativity for BzATP than for ATP in opening the P2Z receptor-operated ion channel.
  3. A rank order of agonist potency of BzATP>ATP=2MeSATP>ATPγS was observed for agonist-stimulated ethidium+ influx, while maximal influxes followed a rank order of BzATP>ATP>2MeSATP>ATPγS.
  4. Preincubation with 30–50 μM oxidized ATP (ox-ATP), an irreversible P2Z inhibitor, reduced the maximal response but did not change the steepness of the Ba2+ influx-response curve produced by BzATP (nH 3.2 and 2.9 for 30 and 50 μM ox-ATP, respectively (n=2)).
  5. ATP (300–1000 μM) added simultaneously with 30 μM BzATP (EC90) inhibited both ethidium+ and Ba2+ fluxes to a maximum of 30–40% relative to the values observed with BzATP alone. Moreover, ATP (300 μM) shifted the concentration-response curve to the right for BzATP-stimulated Ba2+ influx, confirming competition between ATP and BzATP.
  6. KN-62, a new and powerful inhibitor of the lymphocyte P2Z receptor, showed less potency in antagonizing BzATP-mediated fluxes than ATP-induced fluxes when maximal concentrations of both agonists (BzATP, 50 μM; ATP, 500 μM) were used.
  7. These data suggest that the natural ligand, ATP, is a partial agonist for the P2Z receptor while BzATP is a more efficacious agonist. Moreover the competitive studies show that only a single class of P2-receptor (P2Z class) is expressed on human leukaemic lymphocytes.
  相似文献   

7.
  1. ATP (10–100 μM), but not glutamate (100  μM), stimulated the release of plasminogen from microglia in a concentration-dependent manner during a 10 min stimulation. However, neither ATP (100 μM) nor glutamate (100 μM) stimulated the release of NO. A one hour pretreatment with BAPTA-AM (200 μM), which is metabolized in the cytosol to BAPTA (an intracellular Ca2+ chelator), completely inhibited the plasminogen release evoked by ATP (100 μM). The Ca2+ ionophore A23187 induced plasminogen release in a concentration-dependent manner (0.3 μM to 10 μM).
  2. ATP induced a transient increase in the intracellular calcium concentration ([Ca2+]i) in a concentration-dependent manner which was very similar to the ATP-evoked plasminogen release, whereas glutamate (100 μM) had no effect on [Ca2+]i (70 out of 70 cells) in microglial cells. A second application of ATP (100 μM) stimulated an increase in [Ca2+]i similar to that of the first application (21 out of 21 cells).
  3. The ATP-evoked increase in [Ca2+]i was totally dependent on extracellular Ca2+, 2-Methylthio ATP was active (7 out of 7 cells), but α,β-methylene ATP was inactive (7 out of 7 cells) at inducing an increase in [Ca2+]i. Suramin (100 μM) was shown not to inhibit the ATP-evoked increase in [Ca2+]i (20 out of 20 cells). 2′- and 3′-O-(4-Benzoylbenzoyl)-adenosine 5′-triphosphate (BzATP), a selective agonist of P2X7 receptors, evoked a long-lasting increase in [Ca2+]i even at 1 μM, a concentration at which ATP did not evoke the increase. One hour pretreatment with adenosine 5′-triphosphate-2′, 3′-dialdehyde (oxidized ATP, 100 μM), a selective antagonist of P2X7 receptors, blocked the increase in [Ca2+]i induced by ATP (10 and 100 μM).
  4. These data suggest that ATP may transit information from neurones to microglia, resulting in an increase in [Ca2+]i via the ionotropic P2X7 receptor which stimulates the release of plasminogen from the microglia.
  相似文献   

8.
  1. The influence of L-NG-nitro-arginine (L-NOARG, 30 μM) on contractile responses to exogenous noradrenaline was studied in the rat anococcygeus muscle.
  2. Noradrenaline (0.1–100 μM) contracted the muscle in a concentration-dependent manner. L-NOARG (30 μM) had no effect on noradrenaline responses.
  3. Phenoxybenzamine (Pbz 0.1 μM) depressed by 46% (P<0.001) the maximum response and shifted to the right (P<0.001) the E/[A] curve to noradrenaline (pEC50 control: 6.92±0.09; pEC50 Pbz: 5.30±0.10; n=20).
  4. The nested hyperbolic null method of analysing noradrenaline responses after phenoxybenzamine showed that only 0.61% of the receptors need to be occupied to elicit 50% of the maximum response, indicating a very high functional receptor reserve.
  5. Contractile responses to noradrenaline after partial α1-adrenoceptor alkylation with phenoxybenzamine (0.1 μM) were clearly enhanced by L-NOARG.
  6. The potentiating effect of L-NOARG on noradrenaline responses after phenoxybenzamine was reversed by (100 μM) L-arginine but not by (100 μM) D-arginine.
  7. These results indicate that spontaneous release of NO by nitrergic nerves can influence the α1-adrenoceptor-mediated response to exogenous noradrenaline.
  相似文献   

9.
  1. Whole cell patch clamp techniques were used to study the effects of 4030W92 (2,4-diamino-5-(2,3-dichlorophenyl)-6-fluoromethylpyrimidine), a new antihyperalgesic agent, on rat dorsal root ganglion (DRG) neurones.
  2. In small diameter, presumably nociceptive DRG neurones under voltage-clamp, 4030W92 (1–100 μM) produced a concentration-related inhibition of slow tetrodotoxin-resistant Na+ currents (TTXR). From a holding potential (Vh) of −90 mV, currents evoked by test pulses to 0 mV were inhibited by 4030W92 with a mean IC50 value of approximately 103 μM.
  3. The inhibitory effect of 4030W92 on TTXR was both voltage- and use-dependent. Currents evoked from a Vh of −60 mV were inhibited by 4030W92 with a mean IC50 value of 22 μM, which was 5 fold less than the value obtained at −90 mV. Repeated activation of TTXR by a train of depolarizing pulses (5 Hz, 20 ms duration) enhanced the inhibitory effects of 4030W92. These data could be explained by a preferential interaction of the drug with inactivation states of the channel. In support of this hypothesis 4030W92 (30 μM) produced a significant hyperpolarizing shift of 10 mV in the slow inactivation curve for TTXR and markedly slowed the recovery from channel inactivation.
  4. Fast TTX-sensitive Na+ currents (TTXS) were also inhibited by 4030W92 in a voltage-dependent manner. The IC50 values obtained from Vhs of −90 mV and −70 mV were 37 μM and 5 μM, respectively. 4030W92 (30 μM) produced a 13 mV hyperpolarizing shift in the steady-state inactivation curve of TTXS.
  5. High threshold voltage-gated Ca2+ currents were only weakly inhibited by 4030W92. The reduction in peak Ca2+ current amplitude produced by 100 μM 4030W92 was 20±6% (n=6). Low threshold T-type Ca2+ currents were inhibited by 17±8% and 43±3% by concentrations of 4030W92 of 30 μM and 100 μM, respectively (n=6).
  6. Under current clamp, some cells exhibited broad TTX-resistant action potentials whilst others showed fast TTX-sensitive action potentials in response to a depolarizing current injection. In most cells a long duration (800 ms) supramaximal current injection evoked a train of action potentials. 4030W92 (10–30 μM) had little effect on the first spike in the train but produced a concentration-related inhibition of the later spikes. The number of spikes per train was significantly reduced from 9.7±1.5 to 4.2±1.0 and 2.6±1.1 in the presence of 10 μM and 30 μM 4030W92, respectively (n=5).
  7. Thus, 4030W92 is a potent voltage- and use-dependent inhibitor of Na+ channels in sensory neurones. This profile can be explained by a preferential action of the drug on a slow inactivation state of the channel that results in a delayed recovery to the resting state. This state-dependent modulation by 4030W92 of Na+ channels that are important in sensory neurone function may underlie or contribute to the antihyperalgesic profile of this compound observed in vivo.
  相似文献   

10.
  1. Penile small arteries (effective internal lumen diameter of 300–600 μm) were isolated from the horse corpus cavernosum and mounted in microvascular myographs in order to investigate the mechanisms underlying the endothelium-dependent relaxations to acetylcholine (ACh) and bradykinin (BK).
  2. In arteries preconstricted with the thromboxane analogue U46619 (3–30 nM), ACh and BK elicited concentration-dependent relaxations, pD2 and maximal responses being 7.71±0.09 and 91±1% (n=23), and 8.80±0.07 and 89±2% (n=24) for ACh and BK, respectively. These relaxations were abolished by mechanical endothelial cell removal, attenuated by the nitric oxide (NO) synthase (NOS) inhibitor, NG-nitro-L-arginine (L-NOARG, 100 μM) and unchanged by indomethacin (3 μM). However, raising extracellular K+ to concentrations of 20–30 mM significantly inhibited the ACh and BK relaxant responses to 63±4% (P<0.01, n=7) and to 59±4% (P<0.01, n=6), respectively. ACh- and BK-elicited relaxations were abolished in arteries preconstricted with K+ in the presence of 100 μM L-NOARG.
  3. In contrast to the inhibitor of ATP-sensitive K+ channels, the blockers of Ca2+-activated K+ (KCa) channels, charybdotoxin (30 nM) and apamin (0.3 μM), each induced slight but significant rightward shifts of the relaxations to ACh and BK without affecting the maximal responses. Combination of charybdotoxin and apamin did not cause further inhibition of the relaxations compared to either toxin alone. In the presence of L-NOARG (100 μM), combined application of the two toxins resulted in the most effective inhibition of the relaxations to both ACh and BK. Thus, pD2 and maximal responses for ACh and BK were 7.65±0.08 and 98±1%, and 9.17±0.09 and 100±0%, respectively, in controls, and 5.87±0.09 (P<0.05, n=6) and 38±11% (P<0.05, n=6), and 8.09±0.14 (P<0.01, n=6) and 98±1% (n=6), respectively, after combined application of charybdotoxin plus apamin and L-NOARG.
  4. The selective inhibitor of guanylate cyclase, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 5 μM) did not alter the maximal responses to either ACh or BK, but slightly decreased the sensitivity to both agonists, δpD2 being 0.25±0.07 (P<0.05, n=6) and 0.62±0.12 (P<0.01, n=6) for ACh and BK, respectively. Combined application of ODQ and charybdotoxin plus apamin produced further inhibition of the sensitivity to both ACh (δpD2=1.39±0.09, P<0.01, n=6) and BK (1.29±0.11, P<0.01, n=6), compared to either ODQ or charybdotoxin plus apamin alone.
  5. Exogenous nitric oxide (NO) present in acidified solutions of sodium nitrite (NaNO2) and S-nitroso-cysteine (SNC) both concentration-dependently relaxed penile resistance arteries, pD2 and maximal responses being 4.84±0.06 and 82±3% (n=12), and 6.72±0.07 and 85±4% (n=19), respectively. Charybdotoxin displaced to the right the dose-relaxation curves for both NO (δpD2 0.38±0.06, P<0.01, n=6) and SNC (δpD2 0.50±0.10, P<0.01, n=5), whereas apamin only reduced sensitivity (δpD2=0.35±0.12, P<0.05, n=5) and maximum response (65±9%, P<0.05, n=6) to SNC. ODQ shifted to the right the dose-relaxation curves to both NO and SNC. The relaxant responses to either NO or SNC were not further inhibited by a combination of ODQ and charybdotoxin or ODQ and charybdotoxin plus apamin, respectively, compared to either blocker alone.
  6. In the presence of 3 μM phentolamine, 5 μM ouabain contracted penile resistance arteries by 50±6% (n=17) of K-PSS, but did not significantly change the relaxant responses to either ACh, BK or NO. However, in the presence of L-NOARG ouabain reduced the ACh- and BK-elicited relaxation from 94±3% to 16±5% (P<0.0001, n=6), and from 98±2% to 13±3% (P<0.0001, n=5), respectively. Combined application of ODQ and ouabain inhibited the relaxations to NO from 92±2% to 26±3% (P<0.0001, n=6).
  7. The present results demonstrate that the endothelium-dependent relaxations of penile small arteries involve the release of NO and a non-NO non-prostanoid factor(s) which probably hyperpolarize(s) smooth muscle by two different mechanisms: an increased charybdotoxin and apamin-sensitive K+ conductance and an activation of the Na+-K+ATPase. These two mechanisms appear to be independent of guanylate cyclase stimulation, although NO itself can also activate charybdotoxin-sensitive K+ channels and the Na+-K+ pump through both cyclic GMP-dependent and independent mechanisms, respectively.
  相似文献   

11.
  1. Opioid agonists inhibit neurogenic mucus secretion in the airways. The mechanism of the inhibition is unknown but may be via opening of potassium (K+) channels. We studied the effect on neurogenic secretion in ferret trachea in vitro of the OP1 receptor (formerly known as δ opioid receptor) agonist [D-Pen2,5]enkephalin (DPDPE), the OP2 receptor (formely κ) agonist U-50,488H, the OP3 receptor (formerly μ) agonist [D-Ala2, N-Me-Phe, Gly-ol5]enkephalin (DAMGO), the ATP-sensitive K+ (KATP) channel inhibitor glibenclamide, the large conductance calcium activated K+ (BKCa) channel blocker iberiotoxin, the small conductance KCa (SKCa) channel blocker apamin, the KATP channel opener levcromakalim, a putative KATP channel opener RS 91309, and the BKCa channel opener NS 1619. Secretion was quantified by use of 35SO4 as a mucus marker.
  2. Electrical stimulation increased tracheal secretion by up to 40 fold above sham-stimulated levels. DAMGO or DPDPE (10 μM each) significantly inhibited neurogenic secretion by 85% and 77%, respectively, effects which were reversed by naloxone. U-50,488H had no significant inhibitory effect on neurogenic secretion, and none of the opioids had any effect on ACh-induced or [Sar9]substance P-induced secretion.
  3. Inhibition of neurogenic secretion by DAMGO or DPDPE was reversed by iberiotoxin (3 μM) but not by either glibenclamide or apamin (0.1 μM each). Iberiotoxin alone did not affect the neurogenic secretory response.
  4. Levcromakalim, RS 91309 or NS 1619 (3 nM–3 μM) inhibited neurogenic secretion with maximal inhibitions at 3 μM of 68%, 72% and 96%, respectively. Neither levcromakalim nor RS 91309 at any concentration tested significantly inhibited acetylcholine (ACh)-induced secretion, whereas inhibition (60%) was achieved at the highest concentration of NS 1619, a response which was blocked by iberiotoxin.
  5. Inhibition of neurogenic secretion by levcromakalim (3 μM) or RS 91309 (30 nM) was inhibited by glibenclamide but not by iberiotoxin. In contrast, inhibition by NS 1619 (30 nM and 3 μM) was blocked by iberiotoxin but not by glibenclamide.
  6. We conclude that, in ferret trachea in vitro, OP1 or OP3 opioid receptors inhibit neurogenic mucus secretion at a prejunctional site and that the mechanism of the inhibition is via opening of BKCa channels. Direct opening of BKCa channels or KATP channels also inhibits neurogenic mucus secretion. In addition, opening of BKCa channels inhibits ACh-evoked secretion of mucus. Drugs which open BKCa channels may have therapeutic anti-secretory activity in bronchial diseases in which neurogenic mechanisms and mucus hypersecretion are implicated in pathophysiology, for example asthma and chronic bronchitis.
  相似文献   

12.
  1. The mechanisms underlying the midazolam-induced relaxation of the noradrenaline (NA)-contraction were studied by measuring membrane potential, isometric force and intracellular concentration of Ca2+([Ca2+]i) in endothelium-denuded muscle strips from the rabbit mesenteric resistance artery. The actions of midazolam were compared with those of nicardipine, an L-type Ca2+-channel blocker.
  2. Midazolam (30 and 100 μM) did not modify either the resting membrane potential or the membrane depolarization induced by 10 μM NA.
  3. NA (10 μM) produced a phasic, followed by a tonic increase in both [Ca2+]i and force. Midazolam (10–100 μM) did not modify the resting [Ca2+]i, but attenuated the NA-induced phasic and tonic increases in [Ca2+]i and force, in a concentration-dependent manner. In contrast, nicardipine (0.3 μM) attenuated the NA-induced tonic, but not phasic, increases in [Ca2+]i and force.
  4. In Ca2+-free solution containing 2 mM EGTA, NA (10 μM) transiently increased [Ca2+]i and force. Midazolam (10–100 μM), but not nicardipine (0.3 μM), attenuated this NA-induced increase in [Ca2+]i and force, in a concentration-dependent manner. However, midazolam (10 and 30 μM), had no effect on the increases in [Ca2+]i and force induced by 10 mM caffeine.
  5. In ryanodine-treated strips, which have functionally lost the NA-sensitive Ca2+- storage sites, NA slowly increased [Ca2+]i and force. Nicardipine (0.3 μM) did not modify the resting [Ca2+]i but partly attenuated the NA-induced increases in [Ca2+]i and force. In the presence of nicardipine, midazolam (100 μM) lowered the resting [Ca2+]i and further attenuated the remaining NA-induced increases in [Ca2+]i and force.
  6. The [Ca2+]i-force relationship was obtained in ryanodine-treated strips by the application of ascending concentrations of Ca2+ (0.16–2.6 mM) in Ca2+-free solution containing 100 mM K+. NA (10 μM) shifted the [Ca2+]i-force relationship to the left and enhanced the maximum Ca2+-induced force. Under these conditions, whether in the presence or absence of 10 μM NA, midazolam (10 and 30 μM) attenuated the increases in [Ca2+]i and force induced by Ca2+ without changing the [Ca2+]i-force relationship.
  7. It was concluded that, in smooth muscle of the rabbit mesenteric resistance artery, midazolam inhibits the NA-induced contraction through its inhibitory action on NA-induced Ca2+ mobilization. Midazolam attenuates NA-induced Ca2+ influx via its inhibition of both nicardipine-sensitive and -insensitive pathways. Furthermore, midazolam attenuates the NA-induced release of Ca2+ from the storage sites. This effect contributes to the midazolam-induced inhibition of the NA-induced phasic contraction.
  相似文献   

13.
  1. The actions of the cannabinoid receptor antagonist, SR 141716A, were examined in rat isolated mesenteric arteries. At concentrations greater than 3 μM, it caused concentration-dependent, but endothelium-independent, relaxations of both methoxamine- and 60 mM KCl-precontracted vessels.
  2. SR 141716A (at 10 μM, but not at 1 μM) inhibited contractions to Ca2+ in methoxamine-stimulated mesenteric arteries previously depleted of intracellular Ca2+ stores. Neither concentration affected the phasic contractions induced by methoxamine in the absence of extracellular Ca2+.
  3. SR 141716A (10 μM) caused a 130 fold rightward shift in the concentration-response curve to levcromakalim, a K+ channel activator, but had no effect at 1 μM.
  4. SR 141716A (10 μM) attenuated relaxations to NS 1619 (which activates large conductance, Ca2+-activated K+ channels; BKCa). The inhibitory effect of SR 141716A on NS 1619 was not significantly different from, and was not additive with, that caused by a selective BKCa inhibitor, iberiotoxin (100 nM). SR 141716A (1 μM) did not effect NS 1619 relaxation.
  5. SR 141716A (10 μM) had no effect on relaxations to the nitric oxide donor S-nitroso-N-acetylpenicillamine, or relaxations to carbachol in the presence of 25 mM KCl.
  6. The results show that, at concentrations of 10 μM and above, SR 141716A causes endothelium-independent vasorelaxation by inhibition of Ca2+ entry. It also inhibits relaxations mediated by K+ channel activation. This suggests that such concentrations of SR 141716A are not appropriate for investigation of cannabinoid receptor-dependent processes.
  相似文献   

14.
  1. In the guinea-pig isolated vas deferens preparation bathed in Tyrode''s solution, the prostacyclin analogues, cicaprost, TEI-9063, iloprost, taprostene and benzodioxane-prostacyclin, enhanced twitch responses to submaximal electrical field stimulation (20%-EFS). The high potency of cicaprost (EC150=1.3 nM) and the relative potencies of the analogues (equi-effective molar ratios=1.0, 0.85, 1.6, 17 and 82, respectively) suggest the involvement of a prostacyclin (IP-) receptor.
  2. Maximum enhancement induced by cicaprost in 2.5 mM K+ Krebs-Henseleit solution was similar to that in Tyrode solution (2.7 mM K+), but was progressively reduced as the K+ concentration was increased to 3.9, 5.9 and 11.9 mM. There was also a greater tendency for the other prostacyclin analogues to inhibit EFS responses in 5.9 mM standard K+ Krebs-Henseleit solution; this may be attributed to their agonist actions on presynaptic EP3-receptors resulting in inhibition of transmitter release.
  3. The EFS enhancing action of cicaprost was not affected by the α1-adrenoceptor antagonist prazosin (100 and 1000 nM). Cicaprost (20 and 200 nM) did not affect contractile responses of the vas deferens to either ATP (5 μM) or α,β-methylene ATP (1 μM) in the presence of tetrodotoxin (TTX, 100 nM). In addition, enhancement by cicaprost of responses to higher concentrations of ATP (30 and 300 μM) in the absence of TTX, as shown previously by others, was not seen. Prostaglandin E2 (PGE2, 10 nM) and another prostacyclin analogue TEI-3356 (20 nM) enhanced purinoceptor agonist responses. Unexpectedly, TTX (0.1 and 1 μM) partially inhibited contractions elicited by 10–1000 μM ATP; contractions elicited by 1–3 μM ATP were unaffected. Further studies are required to establish whether a pre- or post-synaptic mechanism is involved.
  4. In a separate series of experiments, cicaprost (5–250 nM), TEI-9063 (3–300 nM), 4-aminopyridine (10–100 μM) and tetraethylammonium (100–1000 μM) enhanced both 20%-EFS responses and the accompanying overflow of noradrenaline to a similar extent. In further experiments with the EP1-receptor antagonist AH 6809, TEI-3356 (1.0–100 nM) and the EP3-receptor agonist, sulprostone (0.1–1.0 nM) inhibited both maximal EFS responses and noradrenaline overflow, thus confirming previous reports of the high activity of TEI-3356 at the EP3-receptor. Cicaprost had no significant effect on noradrenaline overflow at 10 and 100 nM, but produced a modest inhibition at 640 nM.
  5. In conclusion, our studies show that prostacyclin analogues (particularly TEI-3356) can inhibit EFS responses of the guinea-pig vas deferens by acting as agonists at presynaptic EP3-receptors. Prostacyclin analogues (particularly cicaprost and TEI-9063) can also enhance EFS responses through activation of IP-receptors. The mechanism of the enhancement has not been rigorously established but from our results we favour a presynaptic action to increase transmitter release.
  相似文献   

15.
  1. The present study has utilized the two electrode voltage-clamp technique to examine the pharmacological profile of a splice variant of the rat orthologue of the 5-hydroxytryptamine type 3A subunit (5-HT3A(b)) heterologously expressed in Xenopus laevis oocytes.
  2. At negative holding potentials, bath applied 5-HT (300 nM–10 μM) evoked a transient, concentration-dependent (EC50=1.1±0.1 μM), inward current. The response reversed in sign at a holding potential of −2.1±1.6 mV.
  3. The response to 5-HT was mimicked by the 5-HT3 receptor selective agonists 2-methyl-5-HT (EC50=4.1±0.2 μM), 1-phenylbiguanide (EC50=3.0±0.1 μM), 3-chlorophenylbiguanide (EC50=140± 10 nM), 3,5-dichlorophenylbiguanide (EC50=14.5±0.4 nM) and 2,5-dichlorophenylbiguanide (EC50= 10.2±0.6 nM). With the exception of 2-methyl-5-HT, all of the agonists tested elicited maximal current responses comparable to those produced by a saturating concentration (10 μM) of 5-HT.
  4. Responses evoked by 5-HT at EC50 were blocked by the 5-HT3 receptor selective antagonist ondansetron (IC50=231±22 pM) and by the less selective agents (+)-tubocurarine (IC50=31.9± 0.01 nM) and cocaine (IC50=2.1±0.2 μM).
  5. The data are discussed in the context of results previously obtained with the human and mouse orthologues of the 5-HT3A subunit. Overall, the study reinforces the conclusion that species differences detected for native 5-HT3 receptors extend to, and appear largely explained by, differences in the properties of homo-oligomeric receptors formed from 5-HT3A subunit orthologues.
  相似文献   

16.
  1. The release of neuronal [3H]acetylcholine (ACh) from isolated human bronchi after labelling with [3H]choline was measured to investigate the effects of prostanoids.
  2. A first period of electrical field stimulation (S1) caused a [3H]ACh release of 320±70 and 200±40 Becquerel (Bq) g−1 in epithelium-denuded and epithelium-containing bronchi respectively (P>0.05). Subsequent periods of electrical stimulation (Sn, n=2, 3, and 4) released less [3H]ACh, i.e. decreasing Sn/S1 values were obtained (0.76±0.09, 0.68±0.07 and 0.40±0.04, respectively).
  3. Cumulative concentrations (1–1000 nM) of EP-receptor agonists like prostaglandin E2, nocloprost, and sulprostone (EP1 and EP3 selective) inhibited evoked [3H]ACh release in a concentration dependent manner with IC50 values between 4–14 nM and maximal inhibition of about 70%.
  4. The inhibition of evoked [3H]ACh release by prostaglandin E2, nocloprost and sulprostone was not affected by the DP-, EP1- and EP2-receptor antagonist AH6809 at a concentration of 3 μM, i.e. a 3–30 times greater concentration than its affinity (pA2 values) at the respective receptors.
  5. Circaprost (IP-receptor agonist; 1–100 nM), iloprost (IP- and EP1-receptor agonist; 10-1000 nM) and U-46619 (TP-receptor agonist; 100–1000 nM) did not significantly affect [3H]ACh release.
  6. Blockade of cyclooxygenase by 3 μM indomethacin did not significantly modulate evoked [3H]ACh release in epithelium-containing and epithelium-denuded bronchi. Likewise, the combined cyclo- and lipoxygenase inhibitor BW-755C (20 μM) did not affect evoked [3H]ACh release.
  7. In conclusion, applied prostanoids appear to inhibit [3H]ACh release in epithelium-denuded human bronchi under the present in vitro conditions, most likely via prejunctional prostanoid receptors of the EP3 subtype.
  相似文献   

17.
  1. Clozapine has recently been claimed to behave as a selective and full agonist at the cloned m4 muscarinic receptor artificially expressed in Chinese hamster ovary (CHO) cells. In the present study we have investigated whether clozapine could activate the rat striatal muscarinic receptors coupled to the inhibition of adenylyl cyclase activity, considered as pharmacologically equivalent to the m4 gene product. In addition, we have examined the effect of the drug on various functional responses following the activation of the cloned m4 receptor expressed in CHO cells.
  2. In rat striatum, clozapine (1 nM–10 μM) caused a slight inhibition of forskolin-stimulated adenylyl cyclase activity, which was not counteracted by 10 μM atropine. On the other hand, clozapine antagonized the inhibitory effect of acetylcholine with a pA2 value of 7.51. Moreover, clozapine (1 μM) failed to inhibit dopamine D1 receptor stimulation of adenylyl cyclase activity, but counteracted the inhibitory effect of carbachol (CCh). Clozapine displaced [3H]-N-methylscopolamine ([3H]-NMS) bound to striatal M4 receptors with a monophasic inhibitory curve and a pKi value of 7.69. The clozapine inhibition was not affected by the addition of guanosine-5′-O-(thio)triphosphate (GTPγS).
  3. In intact CHO cells, clozapine inhibited forskolin-stimulated cyclic AMP accumulation with an EC50 of 31 nM. This effect was antagonized by atropine. CCh produced a biphasic effect on cyclic AMP levels, inhibiting at concentrations up to 1 μM (EC50=50 nM) and stimulating at higher concentrations (EC50=7 μM). Clozapine (0.3–5 μM) antagonized the CCh stimulation of cyclic AMP with a pKi value of 7.47. Similar results were obtained when the adenylyl cyclase activity was assayed in CHO cell membranes.
  4. In CHO cells pretreated with the receptor alkylating agent 1-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (10 μM), the maximal inhibitory effect of clozapine on cyclic AMP formation was markedly reduced, whereas the CCh inhibitory curve was shifted to the right with no change in the maximum.
  5. As in rat striatum, in CHO cell membranes the displacement of [3H]-NMS binding by clozapine yielded a monophasic curve which was not affected by GTPγS.
  6. Clozapine (10 nM–10 μM) had a small stimulant effect (∼20%) on the binding of [35S]-GTPγS to CHO cell membranes, whereas CCh caused a 250% increase of radioligand binding. Moreover, clozapine (50 nM–5 μM) antagonized the CCh-stimulated [35S]-GTPγS binding with a pA2 value of 7.48.
  7. These results show that at the striatal M4 receptors clozapine is a potent and competitive antagonist, whereas at the cloned m4 receptor it elicits both agonist and antagonist effects. Thus, clozapine behaves as a partial agonist, rather than as a full agonist, at the m4 receptor subtype, with intrinsic activity changing as a function of the coupling efficiency of the receptor to effector molecules.
  相似文献   

18.
  1. Cumulative concentration-response curves (CRC) to prostaglandin E1 (PGE1), PGE2, PGD2 and PGF (0.01–30 μM) and to the thromboxane A2 (TXA2) receptor agonist U-46619 (0.01–30 μM) were constructed in human isolated detrusor muscle strips both in basal conditions and during electrical field stimulation.
  2. All the agonists tested contracted the detrusor muscle. The rank order of agonist potency was: PGF>U-46619>PGE2 whereas weak contractile responses were obtained with PGD2 and PGE1. Any of the agonists tested was able to induce a clear plateau of response even at 30 μM.
  3. The selective TXA2 antagonist, GR 32191B (vapiprost), antagonized U-46619-induced contractions with an apparent pKB value of 8.27±0.12 (n=4 for each antagonist concentration). GR 32191B (0.3 μM) did not antagonize the contractile responses to PGF and it was a non-surmountable antagonist of PGE2 (apparent pKB of 7.09±0.04; n=5). The EP receptor antagonist AH 6809 at 10 μM shifted to the right the CRC to U-46619 (apparent pKB value of 5.88±0.04; n=4).
  4. Electrical field stimulation (20 Hz, 70 V, pulse width 0.1 ms, trains of 5 s every 60 s) elicited contractions fully sensitive to TTX (0.3 μM) and atropine (1 μM). U-46619 (0.01–3 μM) potentiated the twitch contraction in a dose-dependent manner and this effect was competitively antagonized by GR 32191B with an estimated pKB of 8.54±0.14 (n=4 for each antagonist concentration). PGF in the range 0.01–10 μM (n=7), but not PGE2 and PGE1 (n=3 for each), also potentiated the twitch contraction of detrusor muscle strips (23.5±0.3% of KCl 100 mM-induced contraction) but this potentiation was unaffected by 0.3 μM GR 32191B (n=5).
  5. Cumulative additions of U-46619 (0.01–30 μM) were without effect on contractions induced by direct smooth muscle excitation (20 Hz, 40 V, 6 ms pulse width, trains of 2 s every 60 s, in the presence of TTX 1 μM; n=3). Moreover, pretreatment of the tissue with 0.3 μM U-46619 did not potentiate the smooth muscle response to 7 μM bethanecol (n=2).
  6. We concluded that TXA2 can induce direct contraction of human isolated urinary bladder through the classical TXA2 receptor. Prostanoid receptors, fully activated by PGE2 and PGF are also present. All these receptors are probably located post-junctionally. The rank order of agonist potency and the fact that GR32191B, but not AH6809, antagonized responses to PGE2 seem to indicate the presence of a new EP receptor subtype. Moreover, we suggest the presence of prejuctional TXA2 and FP receptors, potentiating acetylcholine release from cholinergic nerve terminals.
  相似文献   

19.
  1. Stellettamide A (ST-A), a novel marine toxin isolated from a marine sponge, inhibited high K+(72.7 mM)-induced contraction in the smooth muscle of guinea-pig taenia coli with an IC50 of 88 μM.
  2. In the taenia permeabilized with Triton X-100, ST-A inhibited Ca2+ (3 and 10 μM)-induced contractions with an IC50 of 46 μM for 3 μM Ca2+ and 105 μM for 10 μM Ca2+. In the permeabilized taenia, calyculin-A (300 nM), a potent inhibitor of type-1 and type-2A phosphatases, induced sustained contraction in the absence of Ca2+. ST-A had no effect on this contraction.
  3. ST-A inhibited Mg2+-ATPase activity in native actomyosin prepared from chicken gizzard with an IC50 of 25 μM.
  4. In a reconstituted smooth muscle contractile system containing calmodulin, myosin light chain (MLC) and MLC kinase, ST-A inhibited MLC phosphorylation with an IC50 of 152 μM. The inhibitory effect of ST-A was antagonized by increasing the concentration of calmodulin.
  5. ST-A inhibited calmodulin activity, assessed by Ca2+/calmodulin-dependent enzymes, (Ca2+-Mg2+)-ATPase of erythrocyte membrane, with an IC50 of 100 μM and phosphodiesterase prepared from bovine cardiac muscle with an IC50 of 52 μM. The inhibitory effect on phosphodiesterase activity was antagonized by increasing the calmodulin concentration.
  6. Interaction between ST-A and calmodulin was demonstrated by instantaneous quenching of the intrinsic tyrosine fluorescence of calmodulin by ST-A (3–300 μM). Similar results were obtained in the presence or absence of Ca2+ suggesting that ST-A binds to calmodulin and that Ca2+ is not essential for the binding of ST-A to calmodulin.
  7. These results suggest that ST-A, isolated from marine metabolites, is a novel inhibitor of calmodulin.
  相似文献   

20.
  1. The aim of study was to characterize endothelin (ET)-induced vasodilatation in isolated extrapulmonary rat arteries (EPA) and in intrapulmonary arteries (IPA) preconstricted with 1 μM phenylephrine.
  2. The ET-3 (1 nM–100 nM)- and ET-1 (10 nM–100 nM)-induced transient vasodilatations in EPA were more potent than those in IPA. The vasodilatation induced by ET-3 (100 nM) was larger than that induced by ET-1 (100 nM).
  3. Both the ETB antagonist, BQ788 (3 μM) and or endothelium denudation, but not the ETA antagonist, BQ123 (3 μM), abolished the vasodilatation induced by ET-1 or ET-3 (100 nM each) in EPA and in IPA. The ATP-sensitive K+channel blocker, glibenclamide (20 μM) and the nitric oxide synthase inhibitor, NG-monomethyl-L-arginine (L-NMMA, 1 mM) suppressed the ET-induced vasodilatation in EPA and in IPA.
  4. We conclude that the vasodilatation induced by endothelins is markedly reduced in rat isolated IPA, and suggest that the endothelial ETB-mediated vasodilatation varies depending on rat pulmonary arterial regions. Furthermore, ETB-mediated vasodilatation involves activation of ATP-sensitive K+ channels and of nitric oxide synthase in rat isolated EPA and IPA.
  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号