Time-Dependent Increase in Network Response to Stimulation

Abstract

In vitro neuronal cultures have become a popular method with which to probe network-level neuronal dynamics and phenomena in controlled laboratory settings. One of the key dynamics of interest in these in vitro studies has been the extent to which cultured networks display properties indicative of learning. Here we demonstrate the effects of a high frequency electrical stimulation signal in training cultured networks of cortical neurons. Networks receiving this training signal displayed a time-dependent increase in the response to a low frequency probing stimulation, particularly in the time window of 20–50 ms after stimulation. This increase was found to be statistically significant as compared to control networks that did not receive training. The timing of this increase suggests potentiation of synaptic mechanisms. To further investigate this possibility, we leveraged the powerful Cox statistical connectivity method as previously investigated by our group. This method was used to identify and track changes in network connectivity strength.

Time-dependent Increase in the Network Response to the Stimulation of Neuronal Cell Cultures on Micro-electrode Arrays

“…The results showed a pre-training response within 20 ms post-stimulus, though the range of activity was inconsistent after the first response. However, post-training activity exhibited not only a response within the first 20 ms post-stimulus, as seen during pre-training, but it also exhibited significant activity 30-50 ms post-stimulus ( Figure 14 and Figure 15 ) 15 . There was also a statistically significant correlation of "spike frequency" versus "time after stimulus" and of "spike reliability" versus "time after stimulus."…”

“…Figure 4b shows the baseline noise and the filtered raw extracellular signal during 8 bursts of activity before the sorting procedure. To separate the action potentials from the noise, thresholds for each channel are set to 5 times the standard deviation of the baseline noise and are calculated over a 500 ms window 15 .…”

“…Prior to analysis, recorded spikes for each electrode were sorted offline to distinguish between physiological activity and stimulation artifacts using a k-means algorithm and principle component analysis. Signals that had been identified as physiological responses were grouped together to create a population response at each electrode ( Figure 13 and Figure 9 ) 15 .…”

“…Upon completing the pre-training stimulation, administer a "training" protocol to the networks using the same electrodes as in the probing stimulation. Deliver the high-frequency trains once every 2 s, as described in Hamilton et al 15 ( Figure 7b and Figure 7c ). NOTE: The training signal consists of 40 pulse trains.…”

“…One of the key dynamics of interest in these in vitro studies has been the extent to which cultured networks display properties indicative of learning 7 8 9 10 11 12 13 . The Peixoto Lab previously examined the effects of high-frequency training signals, as described in Ruaro et al 14 , on networks of mouse neurons plated on microelectrode arrays 15 . In these experiments, networks displayed an increase in response following training induced by electrical stimulation.…”