Layer 3 Excitatory and Inhibitory Circuitry in the Prefrontal Cortex: Developmental Trajectories and Alterations in Schizophrenia

Abstract

Convergent evidence suggests that schizophrenia is a disorder of neurodevelopment with alterations in both early and late developmental processes hypothesized to contribute to the disease process. Abnormalities in certain clinical features of schizophrenia, such as working memory impairments, depend on distributed neural circuitry including the dorsolateral prefrontal cortex (DLPFC) and appear to arise during the protracted maturation of this circuitry across childhood and adolescence. In particular, the neural circuitry substrate for working memory in primates involves the coordinated activity of excitatory pyramidal neurons and a specific population of inhibitory gamma-aminobutyric acid neurons (i.e., parvalbumin-containing basket cells) in layer 3 of the DLPFC. Understanding the relationships between the normal development of-and the schizophrenia-associated alterations in-the DLPFC circuitry that subserves working memory could provide new insights into the nature of schizophrenia as a neurodevelopmental disorder. Consequently, we review the following in this article: 1) recent findings regarding alterations of DLPFC layer 3 circuitry in schizophrenia, 2) the developmental refinements in this circuitry that occur during the period when the working memory alterations in schizophrenia appear to arise and progress, and 3) how various adverse environmental exposures could contribute to developmental disturbances of this circuitry in individuals with schizophrenia.

Keywords: Cortical development; Dorsolateral prefrontal cortex; Excitation/inhibition balance; Parvalbumin interneurons; Pyramidal cells; Schizophrenia.

Figure 1

Alterations in DLPFC layer 3 pyramidal cells in schizophrenia. (A) Schematic diagram illustrating the circuitry in DLPFC layer 3 thought to be crucial for generating γ frequency oscillations required for working memory. Reciprocal connections furnished by local axon collaterals of pyramidal cells provide recurrent excitation, whereas excitatory innervation from pyramidal cells to PVBCs generates feedback inhibition. P, pyramidal cell; PVBC, parvalbumin basket cell. (B) Micrographs of Golgi-impregnated basilar dendritic shafts and spines from a healthy comparison subject (left) and a subject with schizophrenia (right) illustrating a marked decrement in the density of dendritic spines in schizophrenia. Adapted from (58). (C) Scatter plot demonstrating a lower mean spine density on DLPFC deep layer 3 pyramidal neurons in schizophrenia subjects relative to both healthy and psychiatrically-ill comparison subjects. (D) CDC42 signaling pathways that regulate the contribution of F-actin to dendritic spine structure in the healthy brain. The activity of CDC42 is inhibited by ARHGDIA that suppresses intrinsic GTPase activity. CDC42-CDC42EP pathway: Activated CDC42 inhibits CDC42EPs, which dissociate the complex of septin filaments consolidated by SEPT7 in the spine neck, enabling the transient influx of second messengers and molecules from the parent dendrite that facilitate F-actin mediated growth of spines. CDC42-PAK-LIMK pathway: CDC42 activates PAK, which activates LIMK, resulting in downstream regulation of cofilin family of actin depolymerizing proteins necessary for modulating F-actin turnover. Green arrows indicate activation, and red blunted lines indicate inhibition of each target. (E) Molecular alterations and predicted functional consequences of altered CDC42 signaling pathway components in schizophrenia. The multiple alterations in CDC42 signaling pathways are predicted to destabilize actin dynamics and produce spine deficits preferentially in DLPFC layer 3 pyramidal cells in schizophrenia. Larger and darker ovals indicate higher mRNA levels in schizophrenia, while smaller and lighter ovals indicate lower mRNA levels in schizophrenia. Increased thickness of arrows and blunted lines indicate predicted higher activity in schizophrenia, while dashed, thin arrows and blunted lines indicate predicted lower activity in schizophrenia relative to the healthy state shown in panel D.

Figure 2

Schematic summary of alterations in PVBC and PVChC inputs to DLPFC pyramidal cells in schizophrenia. (A) PVBC innervation of the soma and proximal dendrites of pyramidal cells provides strong and fast feedback inhibition. (B) In the healthy state, the strength of PVBC inputs is regulated by both pre- and post-synaptic molecules (See text for details). (C) Schizophrenia is associated with lower strength of PVBC-mediated inhibition due to: 1) downregulation of GAD67 mRNA and protein leading to reduced GABA synthesis and less GABA (black dots) in synaptic vesicles; 2) lower levels of PV expression decrease calcium buffering within PVBC terminals (lighter pink shading); 3) upregulation of “OR lowers presynaptic release of GABA; 4) downregulation of postsynaptic GABA_A α1-containing receptors in pyramidal cells which lowers chloride influx and post-synaptic strength of inhibition. Please refer to Supplemental Information for details about PVChCs in D–F. (D) PVChCs innervate the axon initial segments of pyramidal cells. (E) In the healthy state, the strength of PVChC inputs is regulated by both pre- and post-synaptic molecules (See Supplemental Information for details). (F) Schizophrenia might be associated with increased strength of GABA neurotransmission due to: 1) downregulation of GAT1 protein leading to increased spillover of GABA from terminals (depicted as a lighter shade of green on the terminal outline); 2) upregulation of GABA_A α2-containing receptors in pyramidal cells which might increase chloride influx and postsynaptic strength of GABA neurotransmission at the axon initial segment. GAD67, glutamic acid decarboxylase, 67kDa; PVBC, parvalbumin basket cell; μOR, “ opioid receptor; GABA α1, GABA_A receptor α1 subunit; PVChC, parvalbumin chandelier cell; GAT1, GABA membrane transporter 1; GABA α2, GABA_A receptor α2 subunit.

Figure 3

Postnatal development of DLPFC layer 3 pyramidal cells. (A) Age-related alterations in mean spine density on dendrites of layer 3 pyramidal cells in monkey DLPFC (x-axis in weeks). Gray bar indicates the peripubertal period in monkeys that is roughly analogous to adolescence in humans (93;119). Adapted from (91). (B) Age-related increase in AMPA relative to NMDA receptor contribution to excitatory postsynaptic currents (EPSCs) in DLPFC layer 3 pyramidal cells (x-axis in months). Adapted from (96). (C and D) Age-related changes in mRNA expression of molecular determinants of glutamate neurotransmission (AMPA receptor subunit GluR1 and NMDA receptor subunit GluN1) in DLPFC layer 3 pyramidal cells (x-axis in months). Adapted from (97). In panels B-D, age groups not sharing the same letter are significantly different (p<0.05).

Figure 4

Postnatal development of PVBCs and PVChCs in the primate DLPFC. (A) Postpubertal monkey DLPFC multi-labeled for NeuN/AnkG (blue; cell body and axon initial segment (AIS) markers), GABA_A receptor γ2 subunit (red; postsynaptic marker), PV and vGAT (green, presynaptic markers). PV/vGAT boutons adjacent to γ2-IR clusters not associated with AnkG-labeled AIS were classified as PVBC (schematic magnification). (B) PV protein levels in PVBC boutons were significantly higher in postpubertal monkeys (left) but the density of PVBC boutons did not differ between prepubertal and postpubertal monkeys (right). (C) GABA_A receptor α1 subunit mRNA expression increases in a protracted fashion during postnatal development in DLPFC layer 3 pyramidal cells. Please refer to Supplemental Information for details about PVChCs in D–F. (D) Postpubertal monkey section multi-labeled for NeuN/AnkG, γ2, PV, and vGAT as in panel A. PVChC cartridges were classified as PV/vGAT boutons adjacent to γ2-IR clusters within AnkG-labeled AISs (schematic magnification). (E) PV protein levels in PVChC cartridges did not differ between prepubertal and postpubertal monkeys (left) but the density of PVChC boutons/AIS were significantly lower in postpubertal monkeys (right). Panels A, B, D, E adapted from (100). (F) Protracted age-related decrease in mRNA expression for the GABA_A receptor α2 subunit in DLPFC layer 3 pyramidal cells. (G) During postnatal development, the amplitude of inhibitory postsynaptic currents (IPSCs) significantly increases in DLPFC layer 3 pyramidal cells (left), whereas the duration of IPSCs significantly decreases (right). Adapted from (101). In all relevant panels, age groups not sharing the same letter are significantly different (p<0.05).