Metabolism of amines in the isolated perfused mesenteric arterial bed of the rat

Abstract

1 Semicarbazide-sensitive amine oxidase (SSAO) activity has been demonstrated in the isolated mesenteric arterial bed of the rat in vitro by studying the metabolism of benzylamine (Bz) and tyramine (Tyr) added to the perfusing fluid. 2 Pretreatment of rats with (E)-2-(3',4'-dimethoxyphenyl)-3-fluoroallylamine (MDL72145), a potent inhibitor of SSAO in rat mesenteric blood vessels, reduced the amount of metabolites, following the addition of Bz (25 uM) or Tyr (100pM) to the perfusing fluid, by 83% and 52% respec-' tively. Inactivation of monoamine oxidase type A (MAO-A) by the addition of clorgyline (10pM) to the perfusing fluid, had little effect on the appearance of metabolites from Tyr. 3 The presence of 3 UM cocaine in the perfusing fluid increased the amount of metabolites produced from Tyr. 4 The metabolites of Tyr appearing in the perfusion fluid from control preparations were 85% p-hydroxyphenylacetic acid and the remainder consisted of a mixture of phydroxyphenylacetaldehyde and, possibly, p-hydroxyphenylethanol. 5 The metabolism of Tyr by homogenates of the rat mesenteric vascular bed was carried out by SSAO (60%) and MAO-A (40%) with very little contribution from MAO-B. Homogenates from rats pretreated with MDL 72145 showed metabolism of Tyr by MAO-A only. 6 These data indicate that SSAO is capable of metabolizing amines present in the fluid perfusing blood vessels to metabolites that are readily released. Histochemical evidence has shown that whereas MAO-A is present in the mitochondria of smooth muscle cells and nerve endings, SSAO is located in the plasma membrane of the smooth muscle cells. This subcellular distribution may explain the differences found between metabolites released from intact vessels and the metabolism seen in homogenates. The identity of the Tyr metabolizing activity in intact vessels that is resistant to both MDL 72145 and clorgyline remains to be determined. IntroductionRat blood vessels possess an amine oxidase activity capable of metabolising tyramine (Tyr), which has different properties from monoamine oxidase (MAO; EC 1.4.3.4) enzymes (Coquil et al., 1973 Callingham & Barrand 1987, for review). SSAO has been shown to reside in the plasma membrane of vascular smooth muscle cells (Wibo et al., 1980;Lyles & Singh, 1985).Homogenates of blood vessels have been widely used to examine the biochemical properties of this enzyme activity (Lewinsohn et al., 1978;Clarke et al., 1982;Precious et al., 1988;Elliott et al., 1989b).The preferred substrate appears to be benzylamine (Bz) but a number of aromatic and aliphatic amines also have been shown to be substrates. In spite of all the work done with homogenates, this enzyme has not been studied in intact blood vessels of the rat and its physiological significance is poorly understood.Studies with intact rat caudal artery confirm the long held view that neuronal uptake mechanisms are important for the inactivation of noradrenaline. Catechol-O-methyltransferase and extraneuronal uptake make a more significant contribution when the ...

The influence of amine metabolizing enzymes on the pharmacology of tyramine in the isolated perfused mesenteric arterial bed of the rat

“…The experimental protocol for the examination of the low dose of Tyr differed slightly from that previously described (Elliott et al, 1989 Figure 1 Typical responses of the isolated perfused mesenteric arterial bed of the rat to infusion of modified Krebs-Henseleit solution containing 10 lM tyramine (Tyr). The infusion started at 'S' and a total of 20ml was perfused at a flow rate of 2 ml min .…”

“…The method of isolation and perfusion at constant flow (2mlmin-1) of the mesenteric arterial bed of the rat was followed as previously described (Elliott et al, 1989) and the pharmacological responses to the infusion of 20ml of Krebs solution containing 100 gM Tyr were recorded at the same time as the metabolites appearing in the perfusate were measured. The 5 groups of preparations examined for their ability to metabolize Tyr were also evaluated for the pressor response to Tyr.…”

“…This enzyme activity has been called semicarbazide-sensitive amine oxidase (SSAO,EC 1.4.3.6) and its ability to metabolize Tyr added to the fluid perfusing isolated mesenteric blood vessels of the rat has been demonstrated (Elliott et al, 1989). Unlike MAO, which is a mitochondrial enzyme (Greenawalt & Schnaitman, 1970), SSAO resides in the plasma membrane of smooth muscle cells (Wibo et al, 1980;Lyles & Singh, 1985).…”

“…The isolated perfused mesenteric arterial bed of the rat was chosen for this purpose. Preliminary results of this study have been described elsewhere (Elliott & Callingham, 1988). …”

“…Although Tyr is a substrate for MAO-B there is little of this enzyme present in rat mesenteric blood vessels and therefore it does not contribute significantly to the metabolism of this amine in homogenates of this tissue (Coquil et al, 1973;Elliott et al, 1989).…”