Date Published: March 16, 2017
Publisher: Public Library of Science
Author(s): Ghulam Murtaza, Petra Mermer, Anna Goldenberg, Uwe Pfeil, Renate Paddenberg, Nobert Weissmann, Guenter Lochnit, Wolfgang Kummer, James West.
The two-pore domain potassium channel KCNK3 (TASK-1) is expressed in rat and human pulmonary artery smooth muscle cells. There, it is associated with hypoxia-induced signalling, and its dysfunction is linked to pathogenesis of human pulmonary hypertension. We here aimed to determine its role in hypoxic pulmonary vasoconstriction (HPV) in the mouse, and hence the suitability of this model for further mechanistic investigations, using appropriate inhibitors and TASK-1 knockout (KO) mice. RT-PCR revealed expression of TASK-1 mRNA in murine lungs and pre-acinar pulmonary arteries. Protein localization by immunohistochemistry and western blot was unreliable since all antibodies produced labelling also in TASK-1 KO organs/tissues. HPV was investigated by videomorphometric analysis of intra- (inner diameter: 25–40 μm) and pre-acinar pulmonary arteries (inner diameter: 41–60 μm). HPV persisted in TASK-1 KO intra-acinar arteries. Pre-acinar arteries developed initial HPV, but the response faded earlier (after 30 min) in KO vessels. This HPV pattern was grossly mimicked by the TASK-1 inhibitor anandamide in wild-type vessels. Hypoxia-provoked rise in pulmonary arterial pressure (PAP) in isolated ventilated lungs was affected neither by TASK-1 gene deficiency nor by the TASK-1 inhibitor A293. TASK-1 is dispensable for initiating HPV of murine intra-pulmonary arteries, but participates in sustained HPV specifically in pre-acinar arteries. This does not translate into abnormal rise in PAP. While there is compelling evidence that TASK-1 is involved in the pathogenesis of pulmonary arterial hypertension in humans, the mouse does not appear to serve as a suitable model to study the underlying molecular mechanisms.
Alveolar hypoxia induces contraction of pulmonary arteries, a phenomenon known as hypoxic pulmonary vasoconstriction (HPV), resulting in redistribution of blood from poorly to optimally ventilated lung segments. Initially, HPV is a protective response but chronic hypoxia may lead to pulmonary hypertension (PH) . The two-pore domain K+ channel, KCNK3 (potassium channel subfamily K member 3), also known as TASK-1 (TWIK-related acid-sensitive K+ channel-1), has been implicated both in molecular mechanisms of HPV and in pathogenesis of PH. It is acid sensitive and inhibited by anandamide  and A293 . Closure of this channel decreases K+ efflux, causing membrane depolarization with subsequent opening of voltage-dependent Ca2+ channels and increase in intracellular Ca2+. Increased concentration of Ca2+ within smooth muscle cells (SMC) of vessels will cause vasoconstriction . Carotid body glomus cells show marked hypoxia-sensitive TASK activity [5, 6] which is absent in TASK-1 knockout (KO) mice [7, 8], and these mice show a marked reduction of the hypoxia-evoked increase in carotid sinus nerve discharge . These data point toward a contribution of TASK-1 in oxygen-dependent cellular signalling.
We investigated the functional consequences of TASK-1 deficiency and inhibition on pulmonary vascular reactions, which are sensitive to hypoxia in the mouse. These include rise in PAP in isolated perfused lungs and constriction of intra- and pre-acinar arteries [24–26]. The impact of TASK-1 gene deficiency on these hypoxic responses was rather modest in that hypoxic PAP response and intra-acinar HPV were unaffected, while only maintenance of HPV for more than 30 min was impaired in pre-acinar arteries. Similarly, acute inhibition of TASK-1 by anandamide allowed for initial development of HPV in WT pre-acinar vessels but resulted in fading of HPV after 30 min. The slight but significant difference between intra- and pre-acinar arteries correlated with TASK-1 mRNA expression which was detected only in pre- but not in intra-acinar arteries. Attempts to validate mRNA expression data by immunolabelling revealed ambiguous results since, in agreement with previous findings [21, 35, 36], all available antibodies produced staining also in TASK-1 KO tissues. 2-D Gel electrophoresis combined with mass spectrometry identified the immunoreactive protein as being unrelated to TASK-1.