Heterogeneity of release-inhibiting muscarinic autoreceptors in heart atria and urinary bladder: a study with M 2 - and M 4 -receptor-deficient mice

Abstract

Release-inhibiting muscarinic autoreceptors were studied in heart atria and the urinary bladder of NMRI mice, M(2)-receptor-deficient mice, M(4)-receptor-deficient mice, and wildtype mice sharing the genetic background of the knockout animals. Segments of the tissues were preincubated with (3)H-choline and then superfused and stimulated electrically. In atrial segments taken from adult mice and stimulated with 120 pulses at 1 Hz, the muscarinic receptor agonist oxotremorine-M reduced the evoked overflow of tritium. Its concentration-response curves in atria from NMRI, M(2)-wildtype, M(4)-wildtype and M(2)-knockout mice were similar, with maximal inhibition by about 75%. In atria from M(4)-knockout mice, the maximal inhibitory effect of oxotremorine-M was reduced to 57%. The concentration-response curves of oxotremorine-M were shifted to the right by ipratropium, methoctramine and pirenzepine. Methoctramine and pirenzepine were approximately equipotent antagonists in all strains except in M(4)-knockout atria in which methoctramine was more potent than pirenzepine. When atria from adult NMRI mice were stimulated by 360 pulses at 3 Hz, ipratropium increased the evoked overflow of tritium both in the absence and in the presence of physostigmine (0.1 microM). In atria taken from 1-day-old NMRI mice, oxotremorine-M failed to reduce the evoked overflow of tritium. In bladder segments taken from adult mice, superfused with medium containing oxotremorine-M (1 microM), and stimulated by 360 pulses at 3 Hz, ipratropium increased the evoked overflow of tritium. Its concentration-response curves in preparations from NMRI, M(2)-wildtype, M(4)-wildtype and M(2)-knockout mice were similar. There was one exception: ipratropium failed to cause an increase in bladder pieces from M(4)-knockout mice. Methoctramine and pirenzepine also increased the evoked overflow of tritium in all strains except the M(4)-knockout. The two antagonists were approximately equipotent in NMRI, M(4)-wildtype and M(2)-knockout preparations but methoctramine was less potent than pirenzepine in M(2)-wildtype preparations. When bladder pieces from adult NMRI mice were superfused with oxotremorine-M-free medium and stimulated by 360 pulses at 3 Hz, ipratropium increased the evoked overflow of tritium in the presence of physostigmine (0.1 microM) but not in its absence. In bladder segments taken from 1-day-old NMRI mice and superfused with medium containing oxotremorine-M (1 microM), ipratropium increased the evoked overflow of tritium in the same way as in adult tissue. It is concluded that NMRI mice and the two wildtype strains are similar in their muscarinic autoreceptors. In atria, the autoreceptors are heterogeneous. Some are M(4). The non-M(4)-autoreceptors probably are M(2). In the bladder, the autoreceptors are exclusively M(4). In both tissues, the autoreceptors are activated by previously released acetylcholine under appropriate conditions.

Activation of Muscarinic Receptors in Rat Bladder Sensory Pathways Alters Reflex Bladder Activity

“…Sixth, OxoM did not significantly affect the amplitude of the contractions, suggesting that OxoM did not penetrate the urothelium to reach the parasympathetic efferent nerves. These nerves contain presynaptic M 1 and M 2 /M 4 mAChRs that modulate transmitter release (Somogyi and de Groat, 1992;Somogyi et al, 1994;Braverman et al, 1998;Zhou et al, 2002). These data provide strong evidence that intravesical OxoM activates mAChRs located in the bladder sensory pathways including urothelium and afferent nerves.…”

Muscarinic receptors 2 and 5 regulate bitter response of urethral brush cells via negative feedback

“…Autocrine negative feedback loops that operate via M2 receptors are commonly observed in cholinergic neurons, including striatal neurons (23), α‐motoneurons (24), and postganglionic parasympathetic neurons (25). In parasympathetic cholinergic neurons, the M4 receptor can also serve as an autoreceptor that inhibits acetylcholine release, and it is the dominant, if not sole, receptor that confers this function in the mouse urinary bladder (19). Both M2 and M4 receptors typically act via G i/o proteins and the downstream inhibition of adenylate cyclase activity and voltage‐activated Ca 2+ channels (26), which fits to the inhibitory effect of the M2 receptor on the bitter stimulus increase in [Ca 2+ ] i that was observed in the present study.…”

Muscarinic Acetylcholine Receptors in the Urinary Tract

“…Provided that results obtained in mice are also representative for human bladder, this ambiguity may be clarified by the use of M 2 and M 4 receptor knockout (KO) animals. In electrically stimulated bladder segments, preincubated with [ 3 H]choline and superfused with oxotremorine, ipratropium stimulated tritium outflow in wild-type and M 2 receptor KO bladders, but had no effect in M 4 receptor KO bladders (Zhou et al 2002). These studies clearly implicate a key role of M 4 receptors in regulating ACh release from postganglionic cholinergic nerves via a negative feedback mechanism.…”