Signal transduction pathways of muscarinic receptor mediated activation in the newborn and adult mouse urinary bladder.
OBJECTIVE
To study the role of M(2) and M(3) muscarinic receptor subtypes, sources of activator Ca(2+), and mechanisms involved in increased force oscillations in muscarinic contractions in the bladders of newborn and adult mice, as in the adult bladder muscarinic M(3) receptors are considered to mediate the main part of bladder contraction, and this has not been established in the newborn bladder.


METHODS
Bladder preparations from newborn (0-2 days) and adult (10-12 weeks) mice were mounted for in vitro force registration and activated with carbachol and high-K(+) solution in the presence of M(3) (4-DAMP 30 nM) or M(2) (methoctramine, 100 nM) receptor antagonists. Thapsigargin (1 microm) or ryanodine (10 microm) were used to inhibit sarcoplasmic reticulum Ca(2+) release. L-NAME (300 microm) and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 10 microm) were used to inhibit nitric oxide synthase and guanylyl cyclase, respectively. Gap-junction function was inhibited with by 18-beta-glycyrrhetinic acid (18-beta-GA; 0.1-100 microm). Big-conductance (BK) and small-conductance (SK) K(+) channels were inhibited by apamine and charybdotoxin (0.3 microm), respectively.


RESULTS
Concentration-response relations for carbachol in the presence of 4-DAMP and methoctramine showed that M(3) receptors are the main activating pathway also in the newborn bladder. Neither thapsigargin nor ryanodine influenced the muscarinic responses of the newborn and adult bladders. Carbachol-induced contractions were not influenced by L-NAME or ODQ. The 18-beta-GA inhibited carbachol-induced contractions in both newborn and adult tissue in a similar manner. Apamine and charybdotoxin slightly increased the amplitude of the contractile responses.


CONCLUSIONS
These results suggest that in the newborn mouse bladder, as in adult bladders, the M(3) muscarinic receptor subtype is mainly responsible for carbachol-induced contractile responses. The main mechanism for muscarinic receptor-induced activation is influx of Ca(2+) from the extracellular medium, and there seems to be no major contribution of Ca(2+) release from intracellular stores. The phasic contractile activity induced by carbachol in the newborn bladder is not influenced by gap junction inhibition and does not involve SK and BK channels.
Ekman
Mari
M
Department of Experimental Medical Science, Lund University, Lund, Sweden.
Andersson
Karl-Erik
KE
Arner
Anders
A
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000037:antagonists & inhibitors / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D017629:Gap Junctions / Q000502:physiology; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000037:antagonists & inhibitors; D008297:Male; D051379:Mice; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D043585:Receptor, Muscarinic M2 / Q000187:drug effects / Q000502:physiology*; D043587:Receptor, Muscarinic M3 / Q000187:drug effects / Q000502:physiology*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D001743:Urinary Bladder / Q000187:drug effects / Q000254:growth & development / Q000502:physiology*
D004791:Enzyme Inhibitors; D051036:Large-Conductance Calcium-Activated Potassium Channels; D043585:Receptor, Muscarinic M2; D043587:Receptor, Muscarinic M3; D002217:Carbachol; D019001:Nitric Oxide Synthase; D006162:Guanylate Cyclase; D002118:Calcium