Biophysical characteristics of activity-induced sodium signals in central neurons
Final Report Abstract
The maintenance of a low intracellular sodium concentration is the basis for the generation of electrical signaling by neurons. In addition, it provides the energy for a multitude of transport processes at the plasma membrane, among them Na+/Ca2+-exchange (NCX), which uses the electrochemical sodium gradient for the export of calcium. Because of the manifold functional consequences of changes in the sodium gradient, understanding of the determinants and the characteristics of activity-induced sodium signals is indispensable. Because these were virtually unknown and un-explored, it was the goal of the present project to provide such information for central neurons in situ, i.e. in brain tissue. To this end, we used quantitative ratiometric and high-resolution sodium imaging, combined with whole-cell patch clamp in CA1 pyramidal neurons of acute tissue slices of the mouse hippocampus. Numerical simulations were performed based on experimentally determined parameters to test the plausibility of our findings. Our results establish that application of glutamate by its focal UV-uncaging results in somatic inward currents accompanied by spatially restriced, local sodium transients in dendrites and spines due to sodium influx through ionotropic glutamate receptors. Recurrent network activity, on the other hand, evokes global sodium network oscillations, comprising both the neuronal as well as the astrocyte cell population. Under these conditions, we found that astrocytes restrict discharge duration and showed that an intact astrocyte metabolism is critical for the neurons’ capacity to recover from sodium loads during synchronized activity. Moreover, and in line with this, we observed that sodium export from CA1 pyramidal cells upon global sodium increases is largely mediated by NKA and, therefore, also depends on an intact neuronal energy metabolism. For recovery from local dendritic sodium increases, in contrast, diffusion dominates over NKA-mediated extrusion, operating efficiently even during short periods of energy deprivation. While sodium will eventually be extruded by the NKA, its diffusion-based fast dissemination to non-stimulated regions might reduce local energy requirements. Diffusion thus plays an important role in the recovery from local sodium transients. Our work showed that sodium ions redistribute along spiny dendrites with an apparent diffusion coefficient of 160 µm²/s, which is considerably slower than expected from previous work. This suggested that diffusion of sodium between dendrites and spines might also be hindered. Indeed, our results revealed that long spine necks serve as significant diffusional barriers for sodium, resulting in a substantial retention of sodium in spines that experienced direct influx. Numerical simulations replicated these findings and also predicted active sodium extrusion from spine necks, supporting that these can represent significant diffusional barriers for sodium. This surprising new property mediates retention of sodium signals in long-neck spines and protects them from dendritic sodium signalling. The latter will promote a reduction in the driving force for sodium-dependent processes. Preliminary experiments reveal a reversal of Na+/Ca2+-exchange upon intracellular sodium increases, indicating that sodium signalling also shapes intracellular calcium signalling. Taken together, our project thus revealed the spatial and temporal properties of activity-induced sodium signals and provided interesting novel insights into the mechanisms of recovery from sodium loads. Furthermore, it revealed new information on sodium diffusion in neuronal microdomains and uncovered an -hitherto unknown- compartmentalization of sodium in long-neck dendritic spines.
Publications
- (2014): Multi-photon intracellular sodium imaging combined with UV-mediated focal uncaging of glutamate in CA1 pyramidal neurons. J. Vis. Exp. (92), e52038
Kleinhans K, Kafitz KW, Rose CR
(See online at https://doi.org/10.3791/2038) - (2015): Astrocytes restrict discharge duration and neuronal sodium loads during recurrent network activity. Glia 63(6):936-57
Karus C, Mondragão M, Ziemens D & Rose CR
(See online at https://doi.org/10.1002/glia.22793) - (2016): Extrusion versus diffusion: mechanisms for recovery from sodium loads in mouse CA1 pyramidal neurons. J. Physiol. Apr 15
Mondragão MA, Schmidt H, Kleinhans C, Langer J, Kafitz KW & Rose CR
(See online at https://doi.org/10.1113/JP272431)
