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and reproduced the interaction. Co-expression of Ca V 2.2/Ca V ␤ 3 subunits with Slo 27 channels revealed rapid functional coupling. By contrast, extremely rare examples of rapid functional coupling were observed with coexpression of Ca V 1.2/Ca V ␤ 3 and Slo 27 channels. Action potential repolarisation in hippocampal pyramidal neurons was slowed by the N-type channel blocker -conotoxin GVIA, but not by the L-type channel blocker isradipine. These data showed that selective functional coupling between N-type Ca 2+ and BK channels provided rapid activation of BK channels in central neurons.

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To fully explore the potential of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs), efficient methods for storage and shipment of these cells are required. Here, we evaluated the feasibility to cold store monolayers and aggregates of functional CMs obtained from different PSC lines using a fully defined clinical-compatible preservation formulation and investigated the time frame that hPSC-CMs could be subjected to hypothermic storage. We showed that two-dimensional (2D) monolayers of hPSC-CMs can be efficiently stored at 4°C for 3 days without compromising cell viability. However, cell viability decreased when the cold storage interval was extended to 7 days. We demonstrated that hPSC-CMs are more resistant to prolonged hypothermic storage-induced cell injury in three-dimensional aggregates than in 2D monolayers, showing high cell recoveries (>70%) after 7 days of storage. Importantly, hPSC-CMs maintained their typical (ultra)structure, gene and protein expression profile, electrophysiological profiles, and drug responsiveness. STEM CELLS TRANSLATIONAL MEDICINE 2016;5:658-669 SIGNIFICANCEThe applicability of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) in the clinic/ industry is highly dependent on the development of efficient methods for worldwide shipment of these cells. This study established effective clinically compatible strategies for cold (4°C) storage of hPSC-CMs cultured as two-dimensional (2D) monolayers and three-dimensional (3D) aggregates. Cell recovery of 2D monolayers of hPSC-CMs was found to be dependent on the time of storage, and 3D cell aggregates were more resistant to prolonged cold storage than 2D monolayers. Of note, it was demonstrated that 7 days of cold storage did not affect hPSC-CM ultrastructure, phenotype, or function. This study provides important insights into the cold preservation of PSC-CMs that could be valuable in improving global commercial distribution of hPSC-CMs.

SUMMARYIn this paper we describe changes in spectral reflectivity of the light reflectors (iridophores) of the squid Alloteuthis subulata. The spectral changes that can be seen in living squid, can also be brought about by superfusing whole skin preparations with acetylcholine (ACh) (20 μmol l-1) and muscarine (30 μmol l-1) but not nicotine (up to 50 mmol l-1), suggesting that cholinergic muscarinic receptors are involved. Changing the osmolarity of the external solution had no effect on spectral reflectivity. To study the iridophores at the cellular level,iridophores were isolated enzymatically. Lucifer Yellow filled the iridophores uniformly, showing cellular individuality. Isolated iridophore cells were loaded with Fura-2 AM and cytoplasmic Ca2+ was recorded ratiometrically. Intracellular Ca2+ (resting concentration at 66.16 nmol l-1) increased transiently after addition of ACh (50 μmol l-1), muscarine (25 μmol l-1), but not nicotine (up to 5 mmol l-1). Ca2+ also increased when superfused with potassium chloride (10 mmol l-1) and caffeine (2.5 mmol l-1). Hypo- and hyperosmotic solutions had no effects on the cytoplasmic Ca2+. By presenting direct evidence that iridophores are polarised cellular structures containing Ca2+ stores and that they are activated via cholinergic muscarinic receptors, we demonstrate that Ca2+ is involved in the reflectivity changes of the iridophores of A. subulata. Specimens were prepared for transmission electron microscopy. It was found that the orientations of the plates with respect to the skin surface are in good agreement with the expected orientations based on the prediction that the iridophores act as multilayer reflectors.