Computational solution of spike overlapping using data-based subtraction algorithms to resolve synchronous sympathetic nerve discharge

Sympathetic nerves conveying central commands to regulate visceral functions often display activities in synchronous bursts. To understand how individual fibers fire synchronously, we establish “oligofiber recording techniques” to record “several” nerve fiber activities simultaneously, using in vitro splanchnic sympathetic nerve–thoracic spinal cord preparations of neonatal rats as experimental models. While distinct spike potentials were easily recorded from collagenase-dissociated sympathetic fibers, a problem arising from synchronous nerve discharges is a higher incidence of complex waveforms resulted from spike overlapping. Because commercial softwares do not provide an explicit solution for spike overlapping, a series of custom-made LabVIEW programs incorporated with MATLAB scripts was therefore written for spike sorting. Spikes were represented as data points after waveform feature extraction and automatically grouped by k-means clustering followed by principal component analysis (PCA) to verify their waveform homogeneity. For dissimilar waveforms with exceeding Hotelling's T(2) distances from the cluster centroids, a unique data-based subtraction algorithm (SA) was used to determine if they were the complex waveforms resulted from superimposing a spike pattern close to the cluster centroid with the other signals that could be observed in original recordings. In comparisons with commercial software, higher accuracy was achieved by analyses using our algorithms for the synthetic data that contained synchronous spiking and complex waveforms. Moreover, both T(2)-selected and SA-retrieved spikes were combined as unit activities. Quantitative analyses were performed to evaluate if unit activities truly originated from single fibers. We conclude that applications of our programs can help to resolve synchronous sympathetic nerve discharges (SND).