Genetic identification of the central nucleus and other components of the central extended amygdala in chicken during development

Abstract

In mammals, the central extended amygdala shows a highly complex organization, and is essential for animal survival due to its implication in fear responses. However, many aspects of its evolution are still unknown, and this structure is especially poorly understood in birds. The aim of this study was to define the central extended amygdala in chicken, by means of a battery of region-specific transcription factors (Pax6, Islet1, Nkx2.1) and phenotypic markers that characterize these different subdivisions in mammals. Our results allowed the identification of at least six distinct subdivisions in the lateral part of the avian central extended amygdala: (1) capsular central subdivision; (2) a group of intercalated-like cell patches; (3) oval central nucleus; (4) peri-intrapeduncular (peri-INP) island field; (5) perioval zone; and (6) a rostral part of the subpallial extended amygdala. In addition, we identified three subdivisions of the laterodorsal bed nucleus of the stria terminalis (BSTLd) belonging to the medial region of the chicken central extended amygdala complex. Based on their genetic profile, cellular composition and apparent embryonic origin of the cells, we discuss the similarity of these different subdivisions of chicken with different parts of the mouse central amygdala and surrounding cell masses, including the intercalated amygdalar masses and the sublenticular part of the central extended amygdala. Most of the subdivisions include various subpopulations of cells that apparently originate in the dorsal striatal, ventral striatal, pallidal, and preoptic embryonic domains, reaching their final location by either radial or tangential migrations. Similarly to mammals, the central amygdala and BSTLd of chicken project to the hypothalamus, and include different neurons expressing proenkephalin, corticotropin-releasing factor, somatostatin or tyrosine hydroxylase, which may be involved in the control of different aspects of fear/anxiety-related behavior.

Embryonic Origin of the Islet1 and Pax6 Neurons of the Chicken Central Extended Amygdala Using Cell Migration Assays and Relation to Different Neuropeptide-Containing Cells

“…On Western blots of turtle and rat brain samples, it recognizes a single band of about 42 kDa, which corresponds to the molecular weight of the protein . The staining pattern obtained with this antibody in the developing chicken forebrain is identical to that obtained with in situ hybridization [Abellán and Medina, 2009;Vicario et al, 2014]. Moreover, the staining pattern obtained with this antibody in the developing chicken forebrain is highly similar to that obtained in mouse and turtle forebrain Bupesh et al, 2011a].…”

“…Similar to the mouse LGEv, the chicken Stv is characterized by expression of Islet1 in the subventricular and mantle zone during development [Abellán and Medina, 2009;Vicario et al, 2014]. Therefore, we used Islet1 as the main genetic marker to be sure of the location of the cell tracker (to distinguish Stv from Std and pallidum, we also performed comparisons with Pax6, Nkx2.1 and other genetic markers of the chicken embryonic brain collection available in our laboratory from previous studies, such as Abellán and Medina [2009] and Vicario et al [2014]). Following CMFDA application in the Stv, we consistently observed numerous labeled cells migrating to the striatal mantle (about 7-13 cells per 100 × 100 μm .…”

“…As in the mouse LGEd, during development, the chicken Std is characterized by expression of Pax6 in the ventricular/subventricular zones and mantle [Abellán and Medina, 2009;Vicario et al, 2014] and expression of FoxP2 in the mantle [García-Calero and Scharff, 2013;. Therefore, we used Pax6 and FoxP2 as the main genetic markers to be sure of the location of the cell tracker (to distinguish Std from Stv and pallium, we also performed comparisons with Islet1, Tbr1, Lhx9, vGLUT2 and other genetic markers of chicken embryonic brain available from our laboratory from previous studies, such as Abellán and Medina In addition, we also consistently observed subpopulations of Std-derived cells that migrated tangentially to either the pallium ( fig.…”

“…Previous studies showed that the embryonic PO (characterized by ventricular zone and mantle expression of Nkx2.1, Shh and Lhx7/8) is another source of Islet1-expressing cells for the telencephalon [Abellán and Medina, 2009;Vicario et al, 2014]. To know whether any of the Islet1 neurons of the proposed subdivisions of the chicken EAce originate in the embryonic PO, we carried out cell migration assays at E10 (an age when these subdivisions are already visible) and earlier (E7-E8, when most of the cells of this domain generate).…”

“…It was raised against the C-terminal residues 178-349 of rat Islet1/Islet2, produces a comparable staining pattern in rat and chicken brain, and recognizes both Islet1 and Islet2 [Thor et al, 1991;VarelaEchavarría et al, 1996]. In the chicken brain, staining with this antiserum co-localizes with the mRNA distribution of Islet1 protein by in situ hybridization histochemistry [Varela-Echavarría et al, 1996;Abellán and Medina, 2009;Vicario et al, 2014]. Moreover, the staining pattern obtained with this antibody in the developing chicken forebrain is highly similar to that obtained in mice and turtles using either the same [Bupesh et al, 2011a] or a different antibody .…”