Factors inducing endothelium‐dependent relaxation in the guinea‐pig basilar artery as estimated from the actions of haemoglobin

Abstract

1 Factors inducing dilatation of guinea-pig basilar artery were investigated in intact and endothelium-denuded tissues by measurement of isometric tension and by electrophysiological methods. 2 The amplitudes of contractions induced by 9,1 1,epithio-1 1,12-methanothromboxane A2 (STA2) and by high K+ were enhanced by haemoglobin (oxyhaemoglobin, Hb) in a concentrationdependent fashion (above 1 pM). For the high K'-induced contraction, the initial tonic component was enhanced to a greater extent than the secondary phasic component. Mechanical responses evoked by STA2 and by high K' were greater in endothelium-denuded tissues, but Hb (below 10pM) had no effect on them.3 Hb (10pM) had no effect on the contractile proteins as estimated from the actions of Hb on Ca2+-induced contractions in skinned muscle tissues. Further, Hb had no effect on the release of Ca2+ from intracellular stores but it accelerated the Ca2 + accumulation into the sarcoplasmic reticulum as judged from the caffeine-or STA2-induced contraction generated in intact tissues. 4 Acetylcholine (ACh) relaxed tissues that were precontracted by STA2 but Hb prevented this relaxation, in a concentration-dependent fashion. The ACh-induced relaxation was sustained for over 10min in the absence of Hb, but following application of Hb, ACh caused only a transient relaxation. 5 STA2 (up to 100nM) did not modify the resting membrane potential of smooth muscle cells of the basilar artery. ACh (1O pM) caused transient hyperpolarization which was only slightly inhibited by Hb (10pM) whether or not STA2 was present. The hyperpolarization induced by ACh required the presence of endothelial cells. 6 A23187 (0.01-1 pM) relaxed tissues which were precontracted by STA2, in a concentrationdependent fashion but had no effect on the membrane potential. 7 These results suggest that in guinea-pig basilar artery, ACh induces relaxation of tissues that were precontracted by STA2 by causing release of both endothelium-derived relaxing (EDRF) and endothelial dependent hyperpolarizing factor (EDHF) (sustained and initial transient relaxation, respectively), but via different mechanisms. Hb inhibits the former and to a lesser extent, the latter.Since A23187 produced relaxation of pre-contracted tissue but caused no detectable change in the membrane potential, this agent may release EDRF but not EDHF.

Cyclic GMP‐independent relaxation and hyperpolarization with acetylcholine in guinea‐pig coronary artery

“…A similar conclusion has been reached by others using various blockers of the NO pathway (Komori et al, 1988;Chen et al, 1988;Huang et al, 1988;Nishiye et al, 1989;Brayden, 1990;Nagao & Vanhoutte, 1991;Chen et al, 1991;Garland & McPherson, 1992). Since combined L-NMMA and HbO entirely abolished the rise in cyclic GMP elicited with ACh our results also provide direct evidence that the EDHF induced hyperpolarization is independent of cyclic GMP.…”

“…However, in several blood vessels the AChinduced hyperpolarization is not mimicked by NO (Beny & Brunet, 1988;Komori et al, 1988;Brayden, 1990;Brayden et al, 1991;Garland & McPherson, 1992) nor can hyperpolarization be blocked by manoeuvres which antagonize the NO pathway such as oxyhaemoglobin (Komori et al, 1988;Chen et al, 1988;Huang et al, 1988;Nishiye et al, 1989) arginine analogues (Nagao & Vanhoutte, 1991;Chen et al, 1991;Garland & McPherson, 1992), and methylene blue (Komori et al, 1988;Chen et al, 1988;Brayden, 1990;Garland & McPherson, 1992). These observations have led to the proposal that a second factor, distinct from NO, is also released from the endothelium in response to ACh stimulation.…”

EDRF‐mediated dilatation in the rat isolated perfused kidney: a microangiographic study

“…70Mm diameter can be adequately resolved (Griffith et al, 1988). Haemoglobin appears to be a specific inhibitor of EDRF activity at the concentration employed (1 gM) (Martin et al, 1985;Edwards et al, 1986;Kelm et al, 1988;Nishiye et al, 1989 (Cocks & Angus, 1983;Miller & Vanhoutte, 1985;Egleme et al, 1984;Matsuda et al, 1985), there is no evidence that al-adrenoceptors can do so in any species or artery type. The lack of effect of haemoglobin in the absence of a constrictor agonist in this preparation differs from previous experiments with the rabbit isolated ear, in which inhibition of basal EDRF activity induced a small but significant rise in perfusion pressure in the absence of exogeneous constrictor agents and in which haemoglobin therefore unmasked myogenic tone (Griffith et al, 1987).…”

“…Methoxamine hydrochloride was a gift from the Wellcome Foundation Ltd, UK, iohexol was obtained from Nyegaard, UK, and Micropaque was obtained from Nicholas, UK. Haemoglobin, which is a specific inhibitor of EDRF activity in peripheral arteries (Martin et al, 1985;Edwards et al, 1986;Nishiye et al, 1989), was prepared following lysis of red blood cells and purified by gel filtration on a Sephadex G200 column as previously described (Edwards et al, 1986 but, 'paradoxically', resulted in dilatation of RO (centre).…”

Actions of a newly synthesized nitro-compound, E-4701, on rabbit vascular smooth muscles.

“…ACh may release both the endothelium-derived relaxing and hyperpolarizing factors by activation of muscarinic M2 and M1-receptor subtypes distributed on the endothelium, respectively (27-29). In the endothelium-denuded rabbit coronary artery, E-4701 (10 fM) inhibited the depolarization induced by direct muscle stimulation of ACh, yet it had no effect on the hyperpolarization induced by the release of the endothelium-dependent hyperpolarizing factor (29,30) and the depolarization pro voked by high K.…”