Alterations of cortical pyramidal neurons in mice lacking high-affinity nicotinic receptors

Abstract

The neuronal nicotinic acetylcholine receptors (nAChRs) are allosteric membrane proteins involved in multiple cognitive processes, including attention, learning, and memory. The most abundant form of heterooligomeric nAChRs in the brain contains the beta2- and alpha4- subunits and binds nicotinic agonists with high affinity. In the present study, we investigated in the mouse the consequences of the deletion of one of the nAChR components: the beta2-subunit (beta2(-/-)) on the microanatomy of cortical pyramidal cells. Using an intracellular injection method, complete basal dendritic arbors of 650 layer III pyramidal neurons were sampled from seven cortical fields, including primary sensory, motor, and associational areas, in both beta2(-/-) and WT animals. We observed that the pyramidal cell phenotype shows significant quantitative differences among different cortical areas in mutant and WT mice. In WT mice, the density of dendritic spines was rather similar in all cortical fields, except in the prelimbic/infralimbic cortex, where it was significantly higher. In the absence of the beta2-subunit, the most significant reduction in the density of spines took place in this high-order associational field. Our data suggest that the beta2-subunit is involved in the dendritic morphogenesis of pyramidal neurons and, in particular, in the circuits that contribute to the high-order functional connectivity of the cerebral cortex.

Fig. 1.

Low-power photomicrographs of horizontal mouse brain slices. In each cortical area sampled, the layer III neurons were injected with Lucifer yellow and then processed with a light-stable diaminobenzidine reaction product. (A) Lateral view of the right hemisphere displaying some of the cortical fields injected: M2, S1 and V2. (B) Medial view of the left hemisphere showing the injected area of PrL/IL. (C) Higher magnification of injected neurons in the M2 field in A. (D) Photomicrograph showing a neuron injected in M2. High-magnification photomicrographs of horizontally projecting dendrites in M2 (E and F) and the anterior cingulate cortex (G and H). (Scale bar: 1.25 mm from A and B; 117 μm in C; 46 μm in D; 12 μm in E–H.)

Fig. 2.

Graphs showing some the morphological parameters analyzed in WT (A–D) and KO (E–H) mice. Individual points in graphs indicate the average values obtained in each animal examined. Bars indicate mean values and SEMs for six animals sampled.

Fig. 3.

Distribution of the three morphological parameters analyzed as a function of the distance from soma: Sholl analysis (first column), spine density (number of spines per 10 μm; second column) and total number of spines (third column). Data from the WT mice are shown in the first row, whereas the second through fifth rows illustrate the comparison of morphological parameters among cortical areas in WT and KO mice.

Fig. 4.

Schematic drawings, represented by color codes, of the main morphological parameters analyzed in the basal dendritic arbors of pyramidal cells in the lateral (M1, S2, M2, V1, V2, and S1) areas in WT (first column) and KO (second column) mice. Significant differences between the two groups of animals are represented in the third column of the figure (WT-KO). Fields 4, 6, 10 and 11 of frontal cortex correspond to primary motor cortex (M1), secondary motor cortex (M2), frontal association cortex (FrA), dorsal agranular insular cortex (AID) and ventral agranular insular cortex (AIV). Fields 1, 2, 3, 3a and 40 of parietal cortex correspond to primary somatosensory cortex (S1), primary somatosensory barrelfield cortex (S1BF) and secondary somatosensory cortex (S2). Fields 22, 36 and 41 of temporal cortex correspond to primary and secondary auditory (A1 and A2) and association area (TeA) of temporal cortex. Fields 17 and 18 of occipital cortex correspond to primary and secondary visual cortex (V1 and V2). Fields 13 and 14 of insular cortex correspond to granular insular (GI) and posterior agranular insular (AIP) cortices.

Fig. 5.

Schematic drawings, represented by color codes, of the main morphological parameters analyzed in the basal dendritic arbors of pyramidal cells in the medial (PrL/IL) area in WT (first column) and KO (second column) mice. Significant differences between the two groups of animals are represented in the third column of the figure (WT-KO). Fields 24 and 25 of anterior medial cortex correspond to prelimbic (PrL), infralimbic (IL), medial orbital (MO), ventral orbital (VO), frontal association (FrA), secondary motor (M2), area 1 and 2 of cingulate (Cg1 and Cg2) cortices. Fields 29b and 29c of posterior medial cortex correspond to retrosplenial agranular (RSA), anterior retrosplenial granular (RSGa) and cingulate/retrosplenial (Cg/R) cortices. Field 8 of frontal cortex corresponds to primary and secondary motor cortices (M1 and M2) and frontal association cortex (FrA). Fields 17 and 18b of occipital cortex correspond to primary and secondary visual cortices (V1 and V2). Fields 27 and 49 of retrohippocampal cortex correspond to anterior retrosplenial granular (RSGa).