Sensitivity of coherent oscillations in rat hippocampus to AC electric fields

Abstract

The sensitivity of brain tissue to weak extracellular electric fields is important in assessing potential public health risks of extremely low frequency (ELF) fields, and potential roles of endogenous fields in brain function. Here we determine the effect of applied electric fields on membrane potentials and coherent network oscillations. Applied DC electric fields change transmembrane potentials in CA3 pyramidal cell somata by 0.18 mV per V m(-1) applied. AC sinusoidal electric fields have smaller effects on transmembrane potentials: sensitivity drops as an exponential decay function of frequency. At 50 and 60 Hz it is approximately 0.4 that for DC fields. Effects of fields of < or = 16 V m(-1) peak-to-peak (p-p) did not outlast application. Kainic acid (100 nm) induced coherent network oscillations in the beta and gamma bands (15-100 Hz). Applied fields of > or = 6 V m(-1) p-p (2.1 V m(-1) r.m.s.) shifted the gamma peak in the power spectrum to centre on the applied field frequency or a subharmonic. Statistically significant effects on the timing of pyramidal cell firing within the oscillation appeared at distinct thresholds: at 50 Hz, 1 V m(-1) p-p (354 mV m(-1) r.m.s.) had statistically significant effects in 71% of slices, and 0.5 V m(-1) p-p (177 mV m(-1) r.m.s.) in 20%. These threshold fields are consistent with current environmental guidelines. They correspond to changes in somatic potential of approximately 70 microV, below membrane potential noise levels for neurons, demonstrating the emergent properties of neuronal networks can be more sensitive than measurable effects in single neurons.

Figure 1. Effect of applied sinusoidal fields on CA3 pyramidal cells

A, transmembrane somatic potential plotted as a function of the strength of applied AC electric fields, for frequencies of 10–100 Hz (symbols given in key) reveals a near-linear relationship over the range of field strengths used here (means and standard errors of the mean of at least 6 slices). An example for a single neuron is shown in supplemental material Fig. 2B. B, recordings from one slice show that slower fields have bigger effects on transmembrane potential (also evident in the pooled data of A). C, the slope of the relationships plotted in A plotted as a function of frequency reveals an exponential decay relationship: V_TM = 0.046 + 0.252e^−0.044fqn (R² = 0.99; ANOVA P < 0.0001). The open circle to the left shows the plateau response to DC fields.

Figure 2. Optical measurement of membrane potential responses to extracellularly applied electric fields in the CA3 region of the hippocampus

A, hippocampal slice was stained with the voltage sensitive dye, RH-795. The slice was trimmed to remove CA1 and placed between parallel field electrodes that applied current parallel to the somato-dendritic axis of CA3c pyramidal neurons. Optical responses were monitored from a 50 μm × 50 μm region of interest, centred over the pyramidal cell layer of the CA3c region (small square). B, optical signals (b, membrane depolarization decreases fluorescent intensity) from the somatic region of CA3c pyramidal neurons corresponding to transmembrane voltage responses to applied electric fields (a, 75 V m⁻¹ amplitude, or 150 V m⁻¹ p-p for AC fields) Optical record is the average of 100 successive sweeps. C, expansion of optical responses to the applied fields. Responses to hyperpolarizing DC fields (Cb) could be fitted by a mono-exponential decays, in this case with a time constant of 21.2 ms (grey line), comparable to those measured electrically. The depolarizing current (Ca) is complicated by substantial voltage-sensitive currents. AC field application (150 V m⁻¹ p-p; c and d) was less effective at inducing voltage changes than DC fields. D, expansion of transmembrane potential change measured with sharp microelectrodes in response to depolarizing (a) and hyperpolarizing (b) field application (8 V m⁻¹; grey curve indicates exponential fit). In this case the traces are single sweeps, which were used for fitting exponentials.

Figure 3. Comparison of optical and electrophysiological estimates of effects of applied fields

Mean ±s.e.m. of the sensitivity of optically measured membrane potentials to AC fields at 16 and 50 Hz, normalized to that at DC, plotted on the curve relating the electrophysiological estimates of sensitivity to frequency, again normalized to the sensitivity at DC.

Figure 4. Effect of AC fields on kainate-induced gamma oscillations

A, field potential recording from stratum pyramidale. A 10 s duration 50 Hz electric field was applied at the time marked by the ‘50 Hz AC’ bar. The ‘control’ bar indicates the period used for the control power spectrum. B, power spectra obtained by FFT for the control and applied field epochs in A. Note the shift in the peak power and frequency. C, the differences in the means (bars = 95% confidence interval) of the power spectra for five 10 s epochs with and without the applied field, showing the increase in power centred on 25 Hz during the applied field, and the decrease at the baseline gamma frequency, centred just under 30 Hz in this case. D, expanded gamma oscillation with the times of the minima in the applied field marked above; note the small negative waves on each cycle of gamma align with alternate minima of the applied field, a process we will call ‘entrainment’. E, top, ‘raster plot’ of times of the gamma cycle minima 30 ms either side of each minimum in the applied field; successive cycles of the applied field are plotted along the y-axis. Bottom, histogram of all the spikes included in the raster plot.

Figure 5. Raster plots of effects of weak sinusoidal fields

A, a relatively sensitive slice reveals modulation of the timing of spikes during gamma by a 1 V m⁻¹ p-p AC field, both in the raster plot (above) and the histogram (below). The horizontal dashed grey line indicates the mean count (and the dotted lines ± 2 s.d., or 95% confidence interval) for similar histograms generated during control epochs preceding the applications of the fields. B, a different slice fails to reveal such a modulation, both by eye and from the Kolmogorov–Smirnov tests described in the text.

Figure 6. Difference spectra

A, difference spectra for relatively sensitive (upper spectrum) and insensitive (lower) slice exposed to a 0.5 V m⁻¹ p-p 50 Hz field. B, a less sensitive slice shows no clear changes at 1 V m⁻¹ p-p, but progressively more marked effects as the fields increase. Those above 6 V m⁻¹ p-p show increases centred on 25 Hz, corresponding to alternate cycles of the applied field; that for 2 V m⁻¹ p-p reveals a smaller shift in the frequency of the peak change. (Differences in power scaled × 10⁻³ in A, and × 10⁻⁵ in B.)