THE EFFECTS OF pH ON THE AFFINITY OF PIRENZEPINE FOR MUSCARINIC RECEPTORS IN THE GUINEA‐PIG ILEUM AND RAT FUNDUS STRIP

The binding of pirenzepine to digitonin‐solubilized rat myocardial muscarinic acetylcholine receptors has been examined at 4°C. Solubilization produced only small changes in the binding of N‐methylscopolamine and atropine. In contrast to the low affinity binding of pirenzepine found to be present in the membranes, high affinity binding was detected in the soluble preparation. In both preparations, pirenzepine binding was complex. High affinity pirenzepine binding (KD ∼ 3 × 10−8M) to the soluble myocardial receptors could be monitored directly using [3H]‐pirenzepine. [3H]‐pirenzepine‐labelled soluble myocardial receptors have a sedimentation coefficient of 11.1 s. This indicates that [3H]‐pirenzepine binds predominantly to the uncoupled form of the receptor. However, [3H]‐ pirenzepine‐agonist competition experiments indicated that the high affinity pirenzepine binding sites are capable of coupling with a guanosine 5′‐triphosphate (GTP)‐binding protein. Pirenzepine affinities for the soluble myocardial receptors were unaffected by their state of association with the GTP‐binding proteins found in the heart. The equilibrium binding properties of the soluble cortical and myocardial receptors were very similar. However, the binding kinetics of the myocardial receptor were much slower. It appears that the membrane environment can affect the affinity of pirenzepine for the rat myocardial muscarinic receptor. Removal of the constraint by solubilization allows the expression of high affinity pirenzepine binding.

Section: Discussionmentioning

confidence: 99%

The binding of pirenzepine to digitonin‐solubilized muscarinic acetylcholine receptors from the rat myocardium

Birdsall

Hulme

Keen

1986

“…The identification and classification of multiple muscarinic receptors in central and peripheral nervous systems has been the object of several investigations using different procedures (for reviews see, Turbanti et al, 1982;Caulfield & Straugham, 1983). Recently, it was reported that pirenzepine, a 'nonclassical' muscarinic antagonist, discriminates between subtypes of muscarinic receptor; that is, in both functional and binding experiments, pirenzepine was found to have a higher affinity for muscarinic receptors of certain brain areas (see reviews by Hammer & Giachetti, 1982;Birdsall & Hulme, 1983) and of sympathetic ganglia (Brown et al, 1980) than for the muscarinic receptors in the isolated ileum or heart (Brown et al, 1980;Fuder et al, 1982;Barlow & Chan, 1982).…”

Section: Introductionmentioning

confidence: 99%

Autoregulation of acetylcholine release from vagus nerve terminals through activation of muscarinic receptors in the dog trachea

Ito

Yoshitomi

1988

The effects of pirenzepine and gallamine on the membrane and contractile properties of smooth muscle cells and on excitatory neuro‐effector transmission in the dog trachea were investigated by means of microelectrode, double sucrose gap and tension recording methods. Pirenzepine (10−7 m) and gallamine (10−5 m) had no effect on the resting membrane potential or the input resistance of the smooth muscle cells. Pirenzepine (10−10‐10−9 m) and gallamine (10−7 m) enhanced the amplitude of twitch contractions evoked by field stimulation in the combined presence of indomethacin (10−5 m) and propranolol (10−6 m). At higher concentrations pirenzepine (10−8 m) inhibited the twitch contractions in a dose‐dependent manner. Both pirenzepine and gallamine in doses over 10−7 and 10−5 m, respectively, reduced muscle tone. Pirenzepine (10−10‐10−9 m) and gallamine (10−7 m) enhanced the amplitude of excitatory junction potentials (e.j.ps) evoked by field stimulation (single or repetitive stimulation). However, a high concentration of pirenzepine (10−8 m) reduced the amplitude of e.j.ps. In parallel with its action on e.j.ps, pirenzepine (over 10−9 m) reduced the response of smooth muscle cells to acetylcholine (ACh), in a dose‐dependent manner. Gallamine (5 × 10−5 m) markedly enhanced the amplitude of e.j.ps but also reduced the response of muscle cells to ACh. ACh (10−10‐10−9 m) inhibited twitch contractions evoked by field stimulation, with a slight increase of resting tension. Gallamine enhanced the summation of e.j.ps during repetitive field stimulation at a high frequency (20 Hz), but was without effect on the depression phenomena of e.j.ps observed during double stimulus experiments at different time intervals (5–60 s). These results indicate that both pirenzepine and gallamine have dual actions on pre‐ and postjunctional muscarinic receptors in dog tracheal tissue. At low concentrations both agents potentiate excitatory neuro‐effector transmission, presumably due to enhancement of release of ACh from vagal nerve terminals through blockade of a negative auto‐regulatory process activated by endogenous ACh. At higher concentrations, these agents inhibit the response of smooth muscle cells to ACh through post‐junctional muscarinic receptors and relaxation of the muscle tissue occurs.

“…Hammer et al (1980) with a lower affinity for pirenzepine are of the M2 subtype (Hammer & Giachetti, 1982). Based on the selectivity for pirenzepine, the muscarinic receptors in the sympathetic ganglia are classified as the M, subtype (Brown et al, 1980), while those in ileal smooth muscle and the heart are ofthe M2 subtype (Brown et al, 1980;Fuder et al, 1982;Barlow & Chan, 1982). In the guinea-pig enteric nervous system, activation of M,-and M2-receptors produces postsynaptic depolarization and presynaptic inhibition of transmitter release, respectively (North et al, 1985).…”

Section: Introductionmentioning

confidence: 99%

Heterogeneous distribution of muscarinic receptors in the rabbit saphenous artery

Komori

Suzukixys

1987

1 The properties of the muscarinic receptors in the rabbit saphenous artery were determined from electrical and mechanical responses of smooth muscle cells produced by acetylcholine (ACh). The inhibitory action of atropine and pirenzepine on the ACh-induced responses was also studied. 5 The excitatory junction potential (ej.p.) evoked by perivascular nerve stimulation was inhibited by ACh (above 10-8 M). The ACh-induced inhibition of the ej.p. was antagonized by atropine more preferentially than by pirenzepine (the ID5o values were 3 x 10-8M for atropine and 6 x 10-6 M for pirenzepine).6 It is concluded that in the rabbit saphenous artery, two subtypes of muscarinic receptor (M, and M2) are located on the endothelial cells. Stimulation of each subtype releases a different substance, i.e., a hyperpolarizing substance (M,-subtype) or a relaxant substance (M2-subtype). In prejunctional nerve terminals, the muscarinic receptors responsible for inhibiting the release of transmitter substances are of the M2-subtype.