HL-1 cells express an inwardly rectifying K+ current activated via muscarinic receptors comparable to that in mouse atrial myocytes

An inwardly rectifying K^+ current is present in atrial cardiac myocytes that is activated by acetylcholine (I_{KACh}). Physiologically, activation of the current in the SA node is important in slowing the heart rate with increased parasympathetic tone. It is a paradigm for the direct regulation of signaling effectors by the Gβγ G-protein subunit. Many questions have been addressed in heterologous expression systems with less focus on the behaviour in native myocytes partly because of the technical difficulties in undertaking comparable studies in native cells. In this study, we characterise a potassium current in the atrial-derived cell line HL-1. Using an electrophysiological approach, we compare the characteristics of the potassium current with those in native atrial cells and in a HEK cell line expressing the cloned Kir3.1/3.4 channel. The potassium current recorded in HL-1 is inwardly rectifying and activated by the muscarinic agonist carbachol. Carbachol-activated currents were inhibited by pertussis toxin and tertiapin-Q. The basal current was time-dependently increased when GTP was substituted in the patch-clamp pipette by the non-hydrolysable analogue GTPγS. We compared the kinetics of current modulation in HL-1 with those of freshly isolated atrial mouse cardiomyocytes. The current activation and deactivation kinetics in HL-1 cells are comparable to those measured in atrial cardiomyocytes. Using immunofluorescence, we found GIRK4 at the membrane in HL-1 cells. Real-time RT-PCR confirms the presence of mRNA for the main G-protein subunits, as well as for M2 muscarinic and A1 adenosine receptors. The data suggest HL-1 cells are a good model to study IKAch

Settler state apologies and the elusiveness of forgiveness : The purification ritual that does not purify

Peer reviewedPostprin

To degrade or not to degrade:mechanisms and significance of endocytic recycling

Newly endocytosed integral cell surface proteins are typically either directed for degradation or subject to recycling back to the plasma membrane. The sorting of integral cell surface proteins, including signalling receptors, nutrient transporters, ion channels, adhesion molecules and polarity markers, within the endo-lysosomal network for recycling is increasingly recognised as an essential feature in regulating the complexities of cell, tissue and organism-level physiology. Historically, endocytic recycling has been regarded as a relatively passive process, where the majority of internalized integral proteins are recycled via an unspecific sequence-independent “bulk membrane flow” pathway. Recent work has increasingly challenged this view. The discovery of sequence-specific sorting motifs and the identification of cargo adaptors and associated coat complexes has begun to uncover the highly orchestrated nature of endosomal cargo recycling, thereby providing new insight into the function and (patho)physiology of this process