Revisiting enigmatic cortical calretinin-expressing interneurons

Abstract

Cortical calretinin (CR)-expressing interneurons represent a heterogeneous subpopulation of about 10-30% of GABAergic interneurons, which altogether total ca. 12-20% of all cortical neurons. In the rodent neocortex, CR cells display different somatodendritic morphologies ranging from bipolar to multipolar but the bipolar cells and their variations dominate. They are also diverse at the molecular level as they were shown to express numerous neuropeptides in different combinations including vasoactive intestinal polypeptide (VIP), cholecystokinin (CCK), neurokinin B (NKB) corticotrophin releasing factor (CRF), enkephalin (Enk) but also neuropeptide Y (NPY) and somatostatin (SOM) to a lesser extent. CR-expressing interneurons exhibit different firing behaviors such as adapting, bursting or irregular. They mainly originate from the caudal ganglionic eminence (CGE) but a subpopulation also derives from the dorsal part of the medial ganglionic eminence (MGE). Cortical GABAergic CR-expressing interneurons can be divided in two main populations: VIP-bipolar interneurons deriving from the CGE and SOM-Martinotti-like interneurons originating in the dorsal MGE. Although bipolar cells account for the majority of CR-expressing interneurons, the roles they play in cortical neuronal circuits and in the more general metabolic physiology of the brain remained elusive and enigmatic. The aim of this review is, firstly, to provide a comprehensive view of the morphological, molecular and electrophysiological features defining this cell type. We will, secondly, also summarize what is known about their place in the cortical circuit, their modulation by subcortical afferents and the functional roles they might play in neuronal processing and energy metabolism.

Keywords: embryonic and fetal development; neocortex; neocortical circuits; neuroenergetics; neuropeptides.

Figure 1

Embryonic origins and genetic factors affecting the mature fate of cortical interneurons. (A) Diagram showing the subdivisions of the embryonic telencephalon. The three regions where cortical and hippocampal interneurons originate are the medial ganglionic eminence (MGE) (including the dorsal MGE-dMGE), the caudal ganglionic eminence (CGE), and the preoptic area (POA). The lateral ganglionic eminence (LGE) gives rise, amongst other, to basal forebrain neurons. (B) Ball scheme of the major classes of cortical and hippocampal interneurons identified using neurochemical markers and represented depending of their place of origin in the embryonic telencephalon. The MGE generates 50% of all cortical interneurons and includes mainly parvalbumin (PV)-expressing and somatostatin (SOM)-expressing subtypes. In hippocampus, it also includes a large population expressing the neuronal isoform of nitric oxide synthase (nNOS). CGE-derived reelin (Rln)-expressing interneurons represent the neurogliaform cells. In both cortex and hippocampus, almost all CGE-derived interneurons express the type 3 serotonin receptor (5-HT3 R). In contrast with cortex, hippocampal SOM+ interneuron have a dual origin with a significant subset co-expressing 5-HT3 R. VIP; vasoactive intestinal polypeptide, CR, calretinin; CCK, cholecystokinin. (C) Genetic programs controlling neurogenesis, cell commitment, tangential, and radial migration and maturation of cortical interneuron. The subdivision of the neuroepithelium can be identified by combinatorial expression of transcription factors involved at different stages of cortical interneuron development. Some of these factors participate broadly in interneuron development such as Dlx and CoupTF gene families. Some transcription factors are unique to specific domains and/or stages of differentiation: Nkx2-1 defines the MGE and activates a cascade of genes including Lhx6, Sox6, and Satb1; Nkx6-2 and GLI1 are enriched in the dMGE. Prox1 and SP8 are expressed in CGE-derived cortical interneurons at all stages of their development (adapted from Kessaris et al., 2014).

Figure 2

Coronal brain section of a VIPcre/tdTomato/GIN mouse additionally stained for calretinin. Immunostaining of calretinin in a coronal brain section of a VIPcre/tdTomato/GIN mouse shows VIP neurons in red, Martinotti cells in green and calretinin in blue in the primary somatosensory cortex (S1Tr). (A) Low magnification of one hemisphere depicting the hippocampus (HC), the lateral ventricle (LV), the amygdala (Amy), the thalamic nucleus ventralis posteromedialis (VPM), and the hypothalamus (HT); the dashed rectangle marks the selected area of (B); scale bar: 1000 μm. (B) Close-up of the rectangle in (A) in a maximum intensity projection; Roman numerals indicate cortical layers; dashed rectangle marks the selected area of (C–C″); scale bar: 250 μm. Please note that VIP neurons co-localizing CR appear pink whereas Martinotti cells that co-localize CR show a cyan-colored soma. (C) Red channel of the inset in (B), showing the tdTomato signal only; (C′) green channel of the inset in (B), showing the GFP only; (C″) blue channel of the inset in (B), showing the labeled calretinin antibody only; scale bar for (C–C″) 20 μm. Please not that nearly all VIP and the single Martinotti cell are co-localizing CR.

Figure 3

Somatodendritic morphology of CR+ interneurons. For details see text.

Figure 4

Single cell RT-PCR analysis of a rat CR+ interneuron. (A) Current-clamp recording obtained in response to a depolarizing current pulse (300 pA). Note the initial burst followed by irregularly discharged action potentials. (B) Single-cell RT-PCR analysis of the same neuron revealing co-expression of CR, VIP, ChAT, GAD65, and GAD67. (C) Comparison of the biocytin labeling of the recorded neuron (left panel) with the immunostaining of the slice with an antibody against CR (right panel). Note the immunoreactivity of the biocytin-labeled cell (scale bar 40 μm, adapted from Cauli et al., 1997). (D) Intracellular biocytin labeling of another CR+ bipolar cell analyzed by singe cell RT-PCR. This neuron had a vertically oriented dendritic arborization. Pial surface is upward (scale bar, 20 μm).

Figure 5

Cortical and subcortical inputs of CR+ bipolar interneurons. Schematic representation summarizing the different local cortical neurons (colored cells) and the long-range cortical and subcortical axon terminals (colored fibers) targeting CR+ bipolar interneurons (centered blue bipolar cell). Dendrites and axons are depicted by thick and thin lines, respectively. Anatomical synaptic connections are represented by triangles and putative volume transmission by spherical gradients. The local neuronal types targeting bipolar CR+ and their respective neurotransmitters are schematized and color-coded; pyramidal cells (black, glutamate), fast spiking (FS)-PV neurons (red, GABA), Martinotti cells (green, GABA) other CR+ bipolar interneurons (blue, GABA and Enk [enkephalin]). Neurotransmitters of the long-range cortical and subcortical fibers are color-coded (Glu [glutamate; Black], 5-HT [serotonin, orange], ACh [acetylcholine, purple], NA [noradrenaline, gray], DA [dopamine, pink]) and their major origins indicated using the same color code.

Figure 6

Neuronal and non-neuronal targets of CR+ bipolar cells. Schematic representation summarizing the different neuronal and non-neuronal targets of CR+ bipolar interneurons (blue neurons) and Martinotti cells (green neuron). Dendrites and axons are depicted by thick and thin lines, respectively. Anatomical synaptic connections are represented by triangles, axonal perivascular appositions by dots and putative volume transmission by spherical gradients. The cell types targeted by CR+ neurons are schematized and color-coded; FS-PV neurons (red), Martinotti cells (green), pyramidal cells (black), glial cells (gray). Astro-glial coverage of blood vessels is illustrated by a gray-colored abluminal side. The specific action of the multiple neurotransmitters released by CR+ neurons is depicted by their location near the specific target and their origin color-coded (bipolar cell, blue; Martinotti cells, green). VIP, vasoactive intestinal polypeptide; CRF, corticotrophin releasing factor; NKB, neurokinin B; Som, somatostatin. The disinhibitory action of bipolar cells is represented by disynaptic circuits involving a bipolar interneuron, an intermediary interneuron (left: FS-PV cell; right: Som-Martinotti cell) and a pyramidal cell, respectively.