A living cell is a nonequilibrium thermodynamic system where, nevertheless, a notion of local equilibria exists. This notion applies to all micro- and nanoscale aqueous volumes, each containing a large number of molecules. This allows one to define sets of local conditions, including thermodynamic ones; for instance, a defined temperature requires thermodynamic equilibrium by definition. Once such a condition is fulfilled, one can control local variables and their gradients to theoretically describe the thermodynamic state of living systems at the micro- and nanoscale. Performing ultra-local experimental manipulations has become possible thanks to the patch-clamp technique, which controls the cell membrane potential, and fluorescence imaging, which monitors molecular concentrations and their intracellular gradients. However, precise temperature gradient control at the micro- and nanoscales has yet to be reliably realized in a living cell. Here, we present a new methodology - microscale control of a temperature gradient profile in aqueous media by a fully optical Diamond Heater-Thermometer in a plug-and-play fiber configuration combined with the patch-clamp technique. In particular, we demonstrate applications of the combined Diamond Heater-Thermometer-patch-clamp approach for fast, reproducible thermal modulation of ionic current from voltage-gated Nav1.5 sodium channels expressed in HEK293 cells and in freshly isolated ventricular mouse cardiomyocytes. Such an approach of manipulating the ultra-local temperature has the potential to uncover previously inaccessible phenomena in various physiological intracellular processes related to the endogenous nanoscale heat sources, such as open ion channels capable of producing Joule heat.
