During sustained depolarization, voltage-gated Ca²⁺ channels progressively undergo a transition to a nonconducting, inactivated state, preventing Ca²⁺ overload of the cell. This transition can be triggered either by the membrane potential (voltage-dependent inactivation) or by the consecutive entry of Ca²⁺(Ca²⁺-dependent inactivation), depending on the type of Ca²⁺ channel. These two types of inactivation are suspected to arise from distinct underlying mechanisms, relying on specific molecular sequences of the different pore-forming Ca²⁺ channel subunits. Here we report that the voltage-dependent inactivation (of the α_1ACa²⁺ channel) and the Ca²⁺-dependent inactivation (of the α_1C Ca²⁺ channel) are similarly influenced by Ca²⁺ channel β subunits. The same molecular determinants of the β subunit, and therefore the same subunit interactions, influence both types of inactivation. These results strongly suggest that the voltage and the Ca²⁺-dependent transitions leading to channel inactivation use homologous structures of the different α₁ subunits and occur through the same molecular process. A model of inactivation taking into account these new data is presented.

The administration of autologous (recipient‐derived) tolerogenic dendritic cells (ATDCs) is under clinical evaluation. However, the molecular mechanisms by which these cells prolong graft survival in a donor‐specific manner is unknown. Here, we tested mouse ATDCs for their therapeutic potential in a skin transplantation model. ATDC injection in combination with anti‐CD3 treatment induced the accumulation of CD8⁺CD11c⁺ T cells and significantly prolonged allograft survival. TMEM176B is an intracellular protein expressed in ATDCs and initially identified in allograft tolerance. We show that Tmem176b^−/− ATDCs completely failed to trigger both phenomena but recovered their effect when loaded with donor peptides before injection. These results strongly suggested that ATDCs require TMEM176B to cross‐present antigens in a tolerogenic fashion. In agreement with this, Tmem176b^−/− ATDCs specifically failed to cross‐present male antigens or ovalbumin to CD8⁺ T cells. Finally, we observed that a Tmem176b‐dependent cation current controls phagosomal pH, a critical parameter in cross‐presentation. Thus, ATDCs require TMEM176B to cross‐present donor antigens to induce donor‐specific CD8⁺CD11c⁺ T cells with regulatory properties and prolong graft survival.

1 The effects of zinc (Zn²⁺) on excitability and ionic conductances were analysed on RINm5F insulinoma cells under whole-cell and outside-out patch-clamp recording conditions. 2 We found that extracellular application of 10-20 μM Zn²⁺ induced a reversible abolition of Ca²⁺ action potential firing, which was accompanied by an hyperpolarisation of the resting membrane potential. 3 Higher concentrations of Zn²⁺, in the tens to hundreds micromolar range, induced a reversible reduction of voltage-gated Ca²⁺ and, to a lesser extent, K⁺ currents. Low-voltage-activated Ca²⁺ currents were more sensitive to Zn²⁺ block than high voltage-activated Ca²⁺ currents. 4 The Zn²⁺-induced hyperpolarisation arose from a dose-dependent increase in a voltage-independent K⁺ conductance that was pharmacologically identified as an ATP-sensitive K⁺ (K_ATP) conductance. The effect was rapid in onset, readily reversible, voltage independent, and related to intracellular ATP concentration. In the presence of 1 mM intracellular ATP, half-maximal activation of K_ATP channels was obtained with extracellular application of 1.7 μM Zn²⁺. 5 Single channel analysis revealed that extracellular Zn²⁺ increased the K_ATP channel open-state probability with no change in the single channel conductance. 6 Our data support the hypothesis that Zn²⁺ binding to K_ATP protein subunits results in an activation of the channels, therefore regulating the resting membrane potential and decreasing the excitability of RINm5F cells. Taken together, our results suggest that Zn²⁺ can influence insulin secretion in pancreatic β-cells through a negative feedback loop, involving both K_ATP and voltage-gated conductances.