In vivo imaging of calcium and glutamate responses to intracortical microstimulation reveals distinct temporal responses of the neuropil and somatic compartments in layer II/III neurons

Objective: Intracortical microelectrode implants can generate a tissue response hallmarked by glial scarring and neuron cell death within 100-150 mu m of the biomaterial device. Many have proposed that any performance decline in intracortical microstimulation (ICMS) due to this foreign body tissue response could be offset by increasing the stimulation amplitude. The mechanisms of this approach are unclear, however, as there has not been consensus on how increasing amplitude affects the spatial and temporal recruitment patterns of ICMS. Approach: We clarify these unknowns using in vivo two-photon imaging of mice transgenically expressing the calcium sensor GCaMP6s in Thy1 neurons or virally expressing the glutamate sensor iGluSnFr in neurons. Calcium and neurotransmitter activity are tracked in the neuronal somas and neuropil during long-train stimulation in Layer II/III of somatosensory cortex. Main results: Neural calcium activity and glutamate release are dense and strongest within 20-40 mu m around the electrode, falling off with distance from the electrode. Neuronal calcium increases with higher amplitude stimulations. During prolonged stimulation trains, a sub-population of somas fail to maintain calcium activity. Interestingly, neuropil calcium activity is 3-fold less correlated to somatic calcium activity for cells that drop-out during the long stimulation train compared to cells that sustain activity throughout the train. Glutamate release is apparent only within 20 mu m of the electrode and is sustained for at least 10s after cessation of the 15 and 20 mu A stimulation train, but not lower amplitudes. Significance: These results demonstrate that increasing amplitude can increase the radius and intensity of neural recruitment, but it also alters the temporal response of some neurons. Further, dense glutamate release is highest within the first 20 mu m of the electrode site even at high amplitudes, suggesting that there may be spatial limitations to the amplitude parameter space. The glutamate elevation outlasts stimulation, suggesting that high-amplitude stimulation may affect neurotransmitter re-uptake. This ultimately suggests that increasing the amplitude of ICMS device stimulation may fundamentally alter the temporal neural response, which could have implications for using amplitude to improve the ICMS effect or offset the effects of glial scarring.