Initiation of sodium spikelets in basal dendrites of neocortical pyramidal neurons - PubMed (original) (raw)

Initiation of sodium spikelets in basal dendrites of neocortical pyramidal neurons

B A Milojkovic et al. J Membr Biol. 2005 Nov.

Abstract

Cortical information processing relies critically on the processing of electrical signals in pyramidal neurons. Electrical transients mainly arise when excitatory synaptic inputs impinge upon distal dendritic regions. To study the dendritic aspect of synaptic integration one must record electrical signals in distal dendrites. Since thin dendritic branches, such as oblique and basal dendrites, do not support routine glass electrode measurements, we turned our effort towards voltage-sensitive dye recordings. Using the optical imaging approach we found and reported previously that basal dendrites of neocortical pyramidal neurons show an elaborate repertoire of electrical signals, including backpropagating action potentials and glutamate-evoked plateau potentials. Here we report a novel form of electrical signal, qualitatively and quantitatively different from backpropagating action potentials and dendritic plateau potentials. Strong glutamatergic stimulation of an individual basal dendrite is capable of triggering a fast spike, which precedes the dendritic plateau potential. The amplitude of the fast initial spikelet was actually smaller that the amplitude of the backpropagating action potential in the same dendritic segment. Therefore, the fast initial spike was dubbed "spikelet". Both the basal spikelet and plateau potential propagate decrementally towards the cell body, where they are reflected in the somatic whole-cell recordings. The low incidence of basal spikelets in the somatic intracellular recordings and the impact of basal spikelets on soma-axon action potential initiation are discussed.

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Figures

Fig. 1

Fig. 1. AP backpropagation during suprathreshold excitatory stimulation

(A) High resolution composite photomicrograph of a prefrontal cortical pyramidal cell stained with JPW3028. The schematic drawing indicates the position of the glutamate-filled pipette on a basal dendrite 95 µm from the center of the soma. (B) One frame from a movie sequence taken with the data acquisition camera at 1 kHz sampling frequency (frame rate). (C) Same as in B, except 8 groups of pixels (1–8), from 8 regions of interest (ROIs), were selected across the dendritic tree. To identify pixels used for spatial averaging within the ROI, each selected pixel is shown with a tiny optical trace superimposed using the Neuroplex software. The sizes of ROIs selected for display in panels D_–_F were (in pixels): 1 = 22; 2 = 19; 3 = 12; 4 = 10; 5 = 18; 6 = 15; 7 = 19; and 8 = 16 pixels. The distances of ROIs from the center of the soma were (in µm): 1 = 100; 2 = 95; 3 = 45; 4 = 75; 5 = 115; 6 = 165; 7 = 95; and 8 = 110 µm. (D) The optical signal from the glutamate stimulation site (ROI 2) is aligned with the optical signal from the proximal segment of the target dendrite (ROI 3), non-target dendrite (ROI 1) and electrical signal from the soma (ROI 0). An asterisk indicates our failure to detect a fast transient on the rising phase of the dendritic glutamate-evoked plateau potential. (E) Optical signals from the oblique dendrite (ROIs 4_–_6) are aligned with the somatic whole-cell recording. (F) Optical signals from two oblique branches are aligned with the somatic electrical trace. Note that all of the dendritic recordings show five action potentials having similar amplitude.

Fig. 2

Fig. 2. Fast spikelet at the beginning of the dendritic plateau potential

(A) One frame selected from the sequence of frames (movie) capturing the glutamate-evoked plateau potential in the basal dendritic tree. The dark region in the center of the soma is a camera-saturation artifact. Glutamate pipette (schematic drawing) positioned on a basal dendrite; 115 lm from the soma. (B) The actual pixels used for spatial averaging are marked (by different shades of grey) inside each of 6 regions of interest (ROIs). The distances of ROIs from the center of the soma were (in µm): 1 = 100; 2 = 95; 3 = 45; 4 = 75; 5 = 115; and 6 = 165 µm, respectively. (C) Spatially averaged traces are aligned to show the temporal relations between electrical transients in different parts of the dendritic tree. Vertical dashed line marks the timing of the glutamate pulse. (D) The initial fast spikelet (arrow) is present only in dendritic segments distal to the glutamate stimulation site (ROIs 1 and 2). Gaussian low-pass = 90 Hz cutoff.

Fig. 3

Fig. 3. Glutamate-evoked basal spikelets

(A) Recording of the glutamate-evoked membrane potential changes in the cell body (upper) and target dendritic segment 85 µm away from the soma (lower). Intensity of glutamate iontophoretic current (Ig) = 0.9 µA. (B) Same cell, same camera pixels (spatially averaged), as in the previous panel except that the _I_g = 1.0 µA.

Fig. 4

Fig. 4. The signs of basal spikelets in somatic recordings

(A) Composite image of JPW1114 filled neuron. The glutamate pipette was positioned 85 lm from the soma. (B) Whole-cell recording of glutamate-evoked UP state-like depolarization. Scale = 40 mV. (C) The section marked by a rectangle in B is aligned with the dendritic optical recording on a faster time scale, to show that the dendritic fast spikelet (arrowhead) caused an inflection in the rising phase of the somatic depolarization (arrow). The optical signal (b-dend) is the product of 18 spatially averaged pixels from the ROI marked by box in A. Vertical dashed line (AP) marks the timing of the first somatic action potential.

Fig. 5

Fig. 5. Fast prepotentials are the hallmark of fast dendritic spikes

(A) Image of the JPW1114 filled neuron captured by the fast data acquisition camera. The glutamate stimulation site was on a basal dendrite 80 µm from the soma (schematic drawing). (B) Optical signals obtained from three basal dendrites (ROIs _1_–8) are aligned with the somatic whole-cell recording (ROI 0). Each optical signal is the product of 6 spatially averaged pixel outputs from regions of interest marked by numbers 1–8 in A. Arrowhead, fast prepotential. Arrow, dendritic fast spike. Asterisk, uneventful charging curve. (C) Pixel outputs from ROIs 1 and 2 were spatially averaged, digitally filtered with a Gaussian low-pass filter (90 Hz cutoff; lower trace), and aligned with the somatic whole-cell record (upper trace) to show the time course and the temporal relationship between the dendritic and somatic signals. Arrowhead and arrow, same as in B. Double arrowhead, plateau phase of the glutamate-evoked depolarization. Inset: Composite photograph of the neuron shown in A.

Fig. 6

Fig. 6. Initiation of basal spikelets at near threshold glutamate concentrations

(A) Fluorescence image obtained with the data acquisition camera. The schematic drawing indicates the position of the glutamate-filled sharp pipette. (B) Intensity of glutamate iontophoretic current was gradually increased in five steps (five sweeps I – V) from 0.8 to 1.2 µA. Optical signal from the non-target dendrite (ROI 3) is aligned with optical signals from the target dendrite (ROIs 1 & 2) and whole cell recording from the soma (ROI 0), for each stimulus intensity (I_–_V). Arrowhead, fast prepotential in somatic recording. Arrow, basal spikelet in dendritic optical recording. Asterisk, absence of any fast transient. (C) The same as in B III ROI 0, shown here on a faster time base.

Fig. 7

Fig. 7. Synaptically-evoked basal spikelets

(A) High resolution fluorescence image of the layer V pyramidal cell. The schematic drawings indicate the positions of glutamate and synaptic stimulation pipettes. (B) An area marked with a white rectangle was captured by a fast data acquisition camera (80 × 80 pixels). (C) Recordings of synaptically-evoked membrane potential transients in the cell body (ROI 0) and basal dendrites, as indicated by boxes in B. Stimulus intensity = 30 µA. Duration = 200 µs. Frequency = 50 Hz. Stimulation artifacts have been truncated. Although the synaptic stimulation was intended for dendrite 3, a substantial synaptic input was received by dendrite 2; capable of triggering two fast electrical transients (arrows). (D) Same cell, same camera pixels as in C, except that the neuron was stimulated by glutamate iontophoresis (_I_g = 2.3 µA, duration = 5 ms).

Fig. 8

Fig. 8

In the majority of neurons glutamate-evoked plateaus were not accompanied by fast prepotentials. Three glutamate pulses (1 Hz) were applied to a basal dendrite (90 µm from the soma), as indicated in the schematic drawing. The dendritic stimulation site was found using the so called “sniffing for dendrites” technique. To monitor the somatic excitability, three glutamate pulses were preceded by direct current injection (200 pA, 250 ms). Inset: The somatic response to a second glutamate pulse is displayed on a faster time base to show the time course of the plateau depolarization. An asterisk marks the initial phase of the somatic signal, where fast prepotentials are expected.

Fig. 9

Fig. 9. Fast prepotentials in vitro

A single glutamate pulse was applied to a basal dendrite 95 µm from the soma (_I_g, 2.4 µA; duration, 5 ms). One fast spikelet (arrowhead) interrupts the charging curve of the UP state-like depolarization.

Fig. 10

Fig. 10

In the majority of spikelet-producing neurons the spikelet production was unreliable. Glutamate pulses were delivered to the same dendritic segment at an interval of 2–3 seconds. Although the position of the glutamate pipette was kept fixed between trials, fast ~10 mV prepotentials were detected in less than 50% of trials. Left inset: Schematic drawing of the experimental paradigm. Right inset: A signal from trial 2 is displayed on a faster time base to show the details of the fast spikelet (arrowhead). In this and the following figure Arabic numerals indicate the chronological progression of the trials.

Fig. 11

Fig. 11. In some neurons the initiation of dendritic spikelets was very reliable

(A) Glutamate pulses of gradually increasing intensity were applied to the same dendritic segment at 2–3 seconds inter-stimulus interval. Inset: The fast prepotential was triggered when Ig was increased from 0.8 µA (sweep 1) to 0.9 µA (sweep 2). (B) Three sweeps are superimposed and shifted in time to show that an increase in glutamate concentration presented to the basal dendrite resulted in a decrease in spikelet amplitude in the soma. Numbers mark the chronological order of the traces. (C) The peak amplitude of the basal spikelet in the soma versus the intensity of glutamate iontophoretic current. All glutamate pulses were of equal duration (5 ms). The glutamate pipette was 80 µm from the soma.

Fig. 12

Fig. 12. The overall incidence of fast spikelets is relatively low

(A) Of 267 neurons, whose basal dendrites were stimulated with supra-threshold glutamate pulses, only 11 neurons produced fast spikelets in a very regular manner (black, ~100% success rate). 36 neurons produced basal spikelets in a very irregular manner (grey, less than 20% of glut, pulses per cell were successful). (B) In the data pooled from 267 neurons only 9.6% of the total number of glutamate stimulations (345 out of 3589) produced basal spikelets in somatic recordings.

Fig. 13

Fig. 13

Fast spikelets are blocked by TTX. Three glutamate pulses (1 Hz) were applied to a basal dendrite, 90 µm from the soma, before (upper) and after the introduction of 1 µ

m

TTX into the bath (lower trace). All stimulation parameters (location (90 µm), intensity (1.2 µA), duration (5 ms) and frequency (1 Hz)) were kept unchanged; c.i. - direct current injection (200 pA, 250 ms). Inset: A characteristic ~10 mV fast spikelet interrupts the rising phase of the glutamate-evoked plateau depolarization under control conditions (normal ACSF). Arrows mark fast spikelets that follow each glutamate pulse. Vertical dashed lines mark the timing of the glutamate pulses (glut.). Note that TTX blocked somatic action potentials and fast prepotentials, but not sustained plateau depolarizations (lower trace).

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