Action potential waveform variability limits multi-unit separation in freely behaving rats.
Extracellular multi-unit recording is a widely used technique to study spontaneous and evoked neuronal activity in awake behaving animals. These recordings are done using either single-wire or multiwire electrodes such as tetrodes. In this study we have tested the ability of single-wire electrodes to discriminate activity from multiple neurons under conditions of varying noise and neuronal cell density. Using extracellular single-unit recording, coupled with iontophoresis to drive cell activity across a wide dynamic range, we studied spike waveform variability, and explored systematic differences in single-unit spike waveform within and between brain regions as well as the influence of signal-to-noise ratio (SNR) on the similarity of spike waveforms. We also modelled spike misclassification for a range of cell densities based on neuronal recordings obtained at different SNRs. Modelling predictions were confirmed by classifying spike waveforms from multiple cells with various SNRs using a leading commercial spike-sorting system. Our results show that for single-wire recordings, multiple units can only be reliably distinguished under conditions of high recording SNR (≥ 4) and low neuronal density (≈ 20,000/ mm(3)). Physiological and behavioural changes, as well as technical limitations typical of awake animal preparations, reduce the accuracy of single-channel spike classification, resulting in serious classification errors. For SNR <4, the probability of misclassifying spikes approaches 100% in many cases. Our results suggest that in studies where the SNR is low or neuronal density is high, separation of distinct units needs to be evaluated with great caution.
Queensland Brain Institute, The University of Queensland, Brisbane, Queensland, Australia.
Peter
P
Stratton
Allen
A
Cheung
Janet
J
Wiles
Eugene
E
Kiyatkin
Pankaj
P
Sah
François
F
Windels
D000200:Action Potentials; D000818:Animals; D001921:Brain / Q000502:physiology; D007478:Iontophoresis; D008297:Male; D051381:Rats; D020318:Rats, Long-Evans