Morphological and functional reorganization of rat medial prefrontal cortex in neuropathic pain

Abstract

Neuropathic pain is a chronic pain that results from lesion or dysfunction of the nervous system. Depression and cognitive decline are often coupled to chronic pain, suggesting the involvement of cortical areas associated with higher cognitive functions. We investigated layer 2/3 pyramidal neurons in acute slices of the contralateral medial prefrontal cortex (mPFC) in the rat spared nerve injury (SNI) model of neuropathic pain and found morphological and functional differences between the mPFC of SNI and sham-operated animals. Basal, but not apical, dendrites of neurons from SNI rats are longer and have more branches than their counterparts in sham-operated animals; spine density is also selectively increased in basal dendrites of neurons from SNI rats; the morphological changes are accompanied by increased contribution to synaptic currents of the NMDA component. Interestingly, the NMDA/AMPA ratio of the synaptic current elicited in mPFC neurons by afferent fiber stimulation shows linear correlation with the rats' tactile threshold in the injured (but not in the contralateral) paw. Our results not only provide evidence that neuropathic pain leads to rearrangement of the mPFC, which may help defining the cellular basis for cognitive impairments associated with chronic pain, but also show pain-associated morphological changes in the cortex at single neuron level.

Fig. 1.

Topography of the pyramidal neurons investigated. Neurons were patched in slices of the mPFC contralateral to the surgery side. (A) A slice containing a biocytin-filled neuron is shown beside a cortical map (adapted from ref. 42). The markers on the map represent the locations of the neurons included in this study (O = cells from sham-operated rats; X = cells from SNI rats). (B) Photographs of two representative biocytin-filled neurons, one from a SNI and one from a sham-operated rat. (C and D) Summary bar charts reporting the topographical coordinates of the neurons included in the study show that the neuronal populations sampled in the two animal groups were overlapping.

Fig. 2.

Increased NMDA/AMPA ratio in SNI mPFC pyramidal cells. (A and B) Whole-cell recordings in pyramidal neurons from a sham-operated rat (A) and a SNI rat (B). AMPA currents were recorded at −80 mV. The dotted lines represent the 0 current levels. The AMPA-selective blocker CNQX (20 μM) was then added to the bath solution and CNQX-resistant NMDA currents were recorded at +40 mV. (C) Summary of the data obtained from 26 cells of sham-operated and 24 cells of SNI rats. The NMDA/AMPA ratio was 0.5 ± 0.06 in neurons from sham and 0.77 ± 0.1 in cells from SNI animals (P < 0.05, two-sided t test). (D) Plot of the NMDA/AMPA ratio against tactile threshold measured in the rats' injured paws on the same day of the electrophysiological recordings (data from SNI and sham-operated rats are plotted together). Regression analysis showed a linear correlation between NMDA/AMPA ratio and behavior (n = 50, r = 0.31, P < 0.05). (E) Plotting the same data against the tactile threshold in the contralateral paw did not produce any significant correlation (n = 50, r = 0.09, P ≫ 0.05).

Fig. 3.

Basic electrophysiological properties are similar in mPFC pyramidal neurons of sham and SNI rats. One week after the surgery, patch-clamp recordings were obtained from acute slices in the presence of blockers of fast synaptic transmission (3 mM kynurenic acid and 50 μM picrotoxin). (A) Current clamp traces obtained in a slice from a sham (upper trace) and a SNI rat in response to injection of hyperpolarizing and depolarizing current pulses. (B) Summary plot of the resting membrane potential value in pyramidal cells from the two groups of rats. (C and D) Bar charts summarizing the value of input resistance and maximum firing rate in cells from the two groups.

Fig. 4.

SNI rat mPFC pyramidal cells have longer basal dendritic arbor. Camera lucida reconstruction of biocytin-filled layer 2/3 mPFC neurons from a sham-operated rat (A) and a SNI rat (B). (C and D) Bar charts summarizing the value of total length of the basal (C) and apical (D) dendrites in 16 cells from sham and 16 cells from SNI rats. The basal dendrite length of neurons from SNI rats was increased by ≈30% (P < 0.05).

Fig. 5.

Basal dendrites of cells from SNI rats are more complex. (A) Sholl analysis of basal dendrites of layer 2/3 mPFC pyramidal neurons from sham-operated (hollow symbols, 16 cells) and SNI (filled symbols, 16 cells) rats. Note that the large difference between the number of crossings in neurons from SNI and sham-operated rats at each point between 40 and 100 μm from the soma. (B) Similar analysis in apical dendrites (16 cells in each group) failed to detect significant differences at any distance from the soma. (*, P < 0.05, ANOVA for repeated measures).

Fig. 6.

Spine density is increased in basal dendrites of SNI neurons. (A and B) Photographs of a mPFC pyramidal neuron from a sham-operated rat (A) and a SNI rat (B). (Scale bar, 100 μm.) Dendritic segments included in the boxed areas are shown at higher magnification in C and D. (Scale bar, 20 μm.) (E) Bar chart of the spine density in apical and basal dendrites of pyramidal cells from sham operated (open columns) and SNI rats (filled columns). (F) Total number of spines, extrapolated from spine density and dendritic length, in apical and basal dendrites of neurons from sham-operated and SNI rats (*, P < 0.05).