GTP-binding protein involvement in membrane currents evoked by carbachol and histamine in guinea-pig ileal muscle.
1. Single smooth muscle cells obtained by enzymic dispersion of the longitudinal muscle layer of guinea-pig ileum were used for recording membrane currents under whole-cell voltage clamp in response to carbachol (100 microM, unless otherwise stated) or histamine (100 microM) applied extracellularly. 2. At a holding potential of 0 mV, a transient outward current was evoked by carbachol and histamine. Responses to the two agonists were very similar in size and time course to the current response to caffeine (10 mM). The response to carbachol was virtually absent in the presence of histamine, and vice versa. Caffeine was without effect in the presence of either of these agonists. Inclusion of EGTA (10 or 20 mM) in the pipette abolished the responses to carbachol, histamine and caffeine. Thus, the outward current responses were considered to represent opening of Ca(2+)-activated K+ channels in response to a massive release of Ca2+ from the same stores by these three agents. 3. An inward current was evoked by carbachol and histamine, but not by caffeine at a holding potential of -40 mV, which was considered to represent opening of cationic channels. The carbachol-induced inward current was much longer in duration and larger in size than the histamine-induced inward current. 4. Inclusion of GDP beta S (2 mM) in the pipette abolished the inward and outward current responses to histamine, but inhibited only part of those to carbachol. 5. When the holding potential was held at 0 mV with inclusion of GTP gamma S (0.1-1 mM) in the pipette, spontaneous transient outward currents appeared immediately after break-through but disappeared a few minutes later. Under these conditions, caffeine (10 mM) was almost without effect, suggesting that GTP gamma S had released Ca2+ stores. When the holding potential was held at -40 mV and GTP gamma S (0.1 or 0.2 mM) was present in the pipette, an inward current developed a few minutes after break-through. During the GTP gamma S-induced inward current, application of carbachol or histamine produced no further inward current. However, when 0.01 mM-GTP gamma S was included in the pipette solution, carbachol- and histamine-induced inward currents were potentiated. 6. Pretreated with 2-5 micrograms/ml pertussis toxin (PTX) did not change noticeably the outward current responses to carbachol and histamine, but abolished or markedly reduced the inward current responses.(ABSTRACT TRUNCATED AT 400 WORDS)
Komori
S
S
Department of Veterinary Science, Faculty of Agriculture, Gifu University, Japan.
Kawai
M
M
Takewaki
T
T
Ohashi
H
H
D000818:Animals; D002217:Carbachol / Q000494:pharmacology*; D046508:Culture Techniques; D004594:Electrophysiology; D019204:GTP-Binding Proteins / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006168:Guinea Pigs; D006632:Histamine / Q000494:pharmacology*; D015640:Ion Channel Gating / Q000187:drug effects*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D009130:Muscle, Smooth / Q000502:physiology*; D037342:Pertussis Toxin; D011188:Potassium / Q000378:metabolism; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006632:Histamine; D002217:Carbachol; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D011188:Potassium