K_Ca3.1 channels facilitate K⁺ secretion or Na⁺ absorption depending on apical or basolateral P2Y receptor stimulation

Abstract

Non-technical summary The epithelial cells lining the ducts of the human mammary gland are responsible for modifying sodium and potassium concentrations in milk by actively absorbing sodium from and secreting potassium into the ductal fluid. In the present study we show that adenosine triphosphate (ATP) and uridine triphosphate (UTP) can stimulate sodium absorption and potassium secretion by a mechanism that involves increasing intracellular calcium and activation of calcium-dependent potassium channels. We discovered that addition of ATP or UTP to the luminal surface stimulates potassium secretion, whereas addition of the same concentrations to the epithelial surface normally exposed to the blood produces an increase in sodium absorption. These results provide a better understanding of the signalling mechanisms that control the concentrations of sodium and potassium present in milk.Abstract Human mammary epithelial (HME) cells express several P2Y receptor subtypes located in both apical and basolateral membranes. Apical UTP or ATP-γ-S stimulation of monolayers mounted in Ussing chambers evoked a rapid, but transient decrease in short circuit current (I sc ), consistent with activation of an apical K + conductance. In contrast, basolateral P2Y receptor stimulation activated basolateral K + channels and increased transepithelial Na + absorption. Chelating intracellular Ca 2+ using the membrane-permeable compound BAPTA-AM, abolished the effects of purinoceptor activation on I sc . Apical pretreatment with charybdotoxin also blocked the I sc decrease by >90% and similar magnitudes of inhibition were observed with clotrimazole and TRAM-34. In contrast, iberiotoxin and apamin did not block the effects of apical P2Y receptor stimulation. Silencing the expression of K Ca 3.1 produced ∼70% inhibition of mRNA expression and a similar reduction in the effects of apical purinoceptor agonists on I sc . In addition, silencing P2Y 2 receptors reduced the level of P2Y 2 mRNA by 75% and blocked the effects of ATP-γ-S by 65%. These results suggest that P2Y 2 receptors mediate the effects of purinoceptor agonists on K + secretion by regulating the activity of K Ca 3.1 channels expressed in the apical membrane of HME cells. The results also indicate that release of ATP or UTP across the apical or basolateral membrane elicits qualitatively different effects on ion transport that may ultimately determine the [Na + ]/[K + ] composition of fluid within the mammary ductal network.

Functional expression of a Kir2.1-like inwardly rectifying potassium channel in mouse mammary secretory cells

“…Therefore, some of these properties appeared to be similar to those reported for cloned K Ca 3.1 (5,13,15). A K Ca 3.1-like, Ca 2ϩ -activated K ϩ channel has been also shown to be present in both the basolateral and apical membranes of an immortalized cell line derived from human mammary epithelial cells (21,31), which are supposed to have properties of ductal epithelial cell (21). Nevertheless, it remains unknown whether other types of K ϩ channel are expressed in mammary epithelial cells.…”

“…Therefore, K ϩ channels at the basolateral membranes of MS cells would not only mediate a pathway, through which K ions transported into the cells by the Na ϩ -K ϩ -ATPase and Na ϩ -K ϩ -2Cl Ϫ symporter are recycled, but would also hyperpolarize the membrane potential to provide the driving force for the apical Cl Ϫ efflux into the lumen. Likewise, K ϩ channels in the apical membrane would be supportive for apical Cl Ϫ secretion and for apical K ϩ secretion and Na ϩ absorption that might occur in mammary epithelial cells (3,31). In line with these potential roles of K ϩ channels, previous patch-clamp studies have identified a Ca 2ϩactivated K ϩ current in mouse mammary secretory epithelial cells (of acinar cell origin) in primary culture (6,8).…”

Peripheral Nervous System

“…ATP acts synergistically with oxytocin to increase intracellular Ca 2+ in mouse mammary myoepithelial cells (Nakano et al 2001). P2Y 2 receptors appear to be involved (Blaug et al 2003;Palmer et al 2011). It is not clear whether the source of ATP is neural or paracrine.…”

Purinergic Signalling in the Reproductive System

“…In the human mammary epithelial cell line 31EG4, P2Y 2 receptors mediate apical Ca 2+ -dependent Cl − and fluid secretion (Blaug et al, 2003). In immortalized human mammary epithelial cells activation of P2Y 2 receptors induces Ca 2+ -dependent K + secretion in the apical membrane of the cells, and Na + absorption in the basolateral membrane (Palmer et al, 2011). In bovine mammary epithelial cells ATP increased Ca 2+ -mediated H + efflux; pre-treatment with the lactogenic hormones prolactin, dexamethasone and insulin attenuated the ATP-induced Ca 2+ i increase and the increases in H + efflux (Katoh et al, 2001).…”