Possible involvement of dual-pore domain potassium channels in the development of pacemaker activity in guinea-pig trachea

G.I. Purves, I.T. Johnson, G. Edwards & A.H.Weston. School of Biological Sciences, University of Manchester, G.38 Stopford Building, Oxford Road, Manchester, M13 9PT.

Pacemaker activity in tracheal smooth muscle preparations has previously been reported by Skogvall et al. (1999). These workers showed that tracheal smooth muscle preparations developed a characteristic bursting pattern of spontaneous myogenic activity when bathed in physiological salt solutions (PSS) bubbled with 12% O2, to generate an oxygen tension believed to mimic that present under in vivo conditions. In the present study, confirmation of the findings of Skogvall et al., (1999) has been obtained and the possible role of dual-pore domain K+ (K2P) channels in the development of such activity has been investigated.

Male Dunkin Hartley guinea-pigs (350–650g) were euthanized with CO2 and exsanguinated by severing the brachial artery. The trachea was dissected free in physiological saline solution (PSS) at pH 7.4 and cut into preparations each containing six cartilaginous rings. All tracheal ring segments were cleaved opposite the muscular tissue with end tied to a fixed rod, the other connected to an isometric transducer and tension was applied (~0.15g). The PSS was gassed using various mixtures containing O2 levels of 6, 12, 21 and 95%, respectively.

Under these conditions, tracheal segments exposed to either 6%, or 12% O2 routinely developed waves of myogenic activity similar to those described by Skogvall et al., (1999). Treatment with bupivacaine (1-100µM), or anandamide (1-10µM) in the presence of AM251 (1µM), a CB1-selective antagonist, each caused a concentration-dependent abolition of tension waves and initiated a marked increase in resting tension of the tracheal segments.

Using gene-specific primers, segments of tracheal smooth muscle were probed using RT-PCR techniques for the presence of mRNA for the K2P channel α-subunits TASK-1, TWIK-1, TWIK-2, TREK-1, TREK-2, TRAAK and THIK-1, all of which were detected in myocyte-rich extracts. Furthermore, immunohistochemical studies showed a K2P protein distribution consistent with the results of the RT-PCR experiments.

The results of these experiments suggest that K2P channels may play a variety of roles in guinea-pig myocytes. The ability of anandamide and of bupivacaine (both TASK-1 inhibitors) to inhibit spontaneous myogenic waves generated under conditions believed to mimic ‘physiological’ O2 tensions specifically suggests that TASK-1 channels may play a key role in the generation of such activity. Speculatively, the opening of such channels by drugs may represent a novel bronchodilator mechanism worthy of exploitation.

Skogvall, S. et al., (1999). Acta Physiol. Scand., 165, 81-93.