Download 1 - Sur Lab

Figure 1. A Platform for the Patterned Spatiotemporal Stimulation of Neuronal
Networks. (A) Schematic of cortical slice interfaced with a chip for whole-cell recording
and control via stimulator (left) and live image (right). (B) The system can be
interchangeably interfaced to commercial arrays from different vendors, such as MCS
(left, with blowup) and MED (right) to enable multi-site stimulation. (C) Chip placed on
stage. (D) Stimulator box – or circuit diagram (left) and inside view of *custom*
stimulator (right). (E) Example pulse delivered to two pins from the stimulator via multielectrode array. Scale bars: 5 ms and 1 V. (F) Cortical slice integrated with the multielectrode array of different spacings, such as 200 μm (left) and 10 μm (right). (G) Multielectrode array interfaced with healthy pyramidal cells from a cortical slice (left) and a
dissociated culture (right). (H) Simultaneous stimulation of 10 randomly selected pins in
distinct, random patterns. The stimulator and multi-electrode array can be used together
to stimulate each pin individually (I) and in conjunction with other pins at distinct and
precise stimulus intensities (J).
Figure 2. Precise control over neuronal activity using the spatiotemporal stimulator.
(A) A cortical slice is interfaced with a chip, and simultaneous patch-clamp is achieved
on a layer 2/3 pyramidal cell, as visualized at 2.5x. Scale bars: 200 μm. Stimulating a pin
during current clamp near the patched cell results in single action potentials (B), which
are abolished with 1μm TTX (C). Scale bars: 50 ms and 30 mV (B, C). (D) Probability of
eliciting an action potential exhibits all or none behavior with stimulus intensity. (E)
Action potentials can be elicited by stimulating pins close to the patched cell within an
approximate radius determined by stimulus intensity. (F) Maximum effective range of
stimulus as a function of stimulus intensity. (G) Spread of activation on a cortical slice
bulk loaded with Oregon Green BAPTA 1-AM while stimulating distinct pins. Each
square within the 4x4 grid represents the location of the stimulated pin. Action potential
response to stimulation is temporally precise (H) and can be elicited in a random pattern
(I). Scale bars: 1.5 s and 4 mV (H). Electrical stimuli are indicated by blue arrows.
Figure 3. Precise control over synaptic transmission. (A) A cortical slice is interfaced
with a chip, and a layer 2/3 pyramidal cell is patched in voltage clamp, as visualized at
2.5x. Scale bars: 200 μm. (B) AMPA-mediated EPSCs can be elicited by stimulating pins
away from patched cell at -60 mV in voltage clamp (top) and abolished with 1 μm NBQX
(bottom). Scale bars: 80 ms and 50 pA. (C) Synaptic transmission can be engaged to
different degrees depending on stimulation intensity. Scale bars: 80 ms and 50 pA. (D)
Distinct EPSCs, presumably arising from distinct subpopulations of cells, can be evoked
by different stimulus locations. Numbers correspond to the row and column of the pin
location. Scale bars: 80ms and 30 pA. (E) Mapped visualization of distinct EPSCs
elicited by stimulation of each pin at 4.5 V, indicating heterogeneous responses from
across the cortex. Scale bars: 200 μm.
Figure 4. Evoked synaptic plasticity at single pathway. (A) A cortical slice is
interfaced with a multi-electrode array; a pyramidal cell is patched, and two distinct pins,
inputs A and B, are identified that elicit synaptic transmission (3-4 V, 0.5 ms), visualized
at 2.5x (A) and recorded at -60 mV in voltage clamp (B). (C) Potentiation can be
achieved by applying tetanic to input A (top), and subsequently to input B (bottom). (D)
Average EPSC traces before and after LTP. (E) Average EPSC amplitude for each input
before and after LTP, indicating a significant increase in EPSC amplitude following
tetanic LTP stimulation.
Figure 5. Selective association of entrained pathways via spatiotemporal
programming. (A) Sample traces elicited in current-clamp from two spatially disparate
inputs before and after stimulation in a close temporal sequence. (B) Mapped
visualization of all pin responses before and after association of two chosen pins. Change
in response is specific to pin targeted by associative programming (far right). (C)
Repetition of associative multi-synaptic plasticity across multiple slices. Inputs are
mediated by AMPA transmission and abolished by 1 μm NBQX (top right).
Figure 6. Sequence “ABCDEFG” learning, by repeated fast-timed activation of
nearby cortical regions. A cortical slice is interfaced with a chip, and a pyramidal cell is
patched, as visualized at 2.5x (A) and 40x (B), and as a schematic (D). (C) A whole cell
recording of the patched cell while a “moving bar” stimulus of 2-4V per pin is delivered
to consecutive columns of pins sweeping across the cortex from left to right at 80ms
intervals between columns. (E) Response to the bar is measured before and after a higherfrequency repetition of the bar’s movement. Mapped visualization of response to
“moving bar” stimulus (F) before and (G) after high-frequency repetition. (H) Average
amplitude of response to “moving bar” stimulus before and after high-frequency
repetition as a function of left-to-right bar position.
Figure 7. Using the multi-electrode array to change dynamics at population level.
(A) A system view, illuminating fluorescence using 488 nm light.(B) Layer 2/3 of a
cortical slice bulk loaded with Oregon Green BAPTA 1-AM, as visualized at 40x. (C)
Actual cortical slice interfaced with a chip, as visualized at 2.5x for experimentation. (D)
Cortical slice from (C) has been bulk loaded. Circles, which are areas around each
electrode, depict the regions of interest used during image analysis to quantify the
magnitude of the calcium response about a given electrode. (E) Sample calcium transient
before sequence training (left). Heat maps of responses measured for each given ROI by
stimulating each of the 12 pins. Each box within the 2x6 grid represents measurements
within a single ROI at that spatial position and is a spatial color map depicting the
efficacy of stimulating each site while recording in the box’s ROI (right).(F) Sample
calcium transients (left) and heat maps of responses after training (right). Most ROIs
show larger responses after training, as seen in blue traces in (E) and (F), and some ROIs
even show responses where there previously were none, as seen in green traces in (E) and
(F).