Molecular phenotyping of transient postnatal tyrosine hydroxylase neurons in the rat bed nucleus of the stria terminalis

Abstract

The bed nucleus of the stria terminalis (BNST) is a complex integrative centre in the forebrain, composed of multiple sub-nuclei, each with discrete populations of neurons. Progress in understanding BNST function, both in the adult and during postnatal maturation, is dependent upon a more complete characterization of neuronal phenotypes in the BNST. The aim of the current study was to define the molecular phenotype of one postnatal BNST neuronal population, in order to identify molecular factors that may underlie both (protein marker-related) immaturity, and secondly, the transience of this phenotype. This BNST population was originally identified by high, but transient expression of the EGR1 transcription factor (TF) in postnatal rat lateral intermediate BNST (BNSTLI). The current results confirm a high level of Egr1 activation in postnatal day 10 (PN10) male BNSTLI that is lost at PN40, and now demonstrate a similar pattern of transient activation in female brains. Apparent cellular immaturity in this population, as indicated by low levels of the adult neuronal marker NeuN/RBFOX3, was found to be uncorrelated with both key neuronal regulator protein expression (SOX2 and REST), and also RBFOX2 protein levels. The BNSTLI neurons have a partial catecholaminergic phenotype (tyrosine hydroxylase-positive/dopa decarboxylase-negative; TH+ve/DDC-ve) that is lost at PN40. In contrast, the co-expressed neuropeptide, somatostatin, is maintained, albeit at lower levels, at PN40. The transcriptional basis of the transient and partial catecholaminergic phenotype was investigated by analysing TFs known to maintain adult dopaminergic (TH+ve/DDC+ve) neuronal phenotypes. The BNSTLI neurons were shown to lack forkhead TFs including FOXA1, FOXA2 and FOXO1. In addition, the BNSTLI neurons had low, primarily cytoplasmic, expression of NR4A2/NURR1, an orphan nuclear receptor that is critical for adult maintenance of midbrain dopamine neurons. These results detail the molecular features of an immature neuronal phenotype, and reveal TF deficiencies that may underlie postnatal transience of the phenotype.

Chemogenetic manipulation of the bed nucleus of the stria terminalis counteracts social behavioral deficits induced by early life stress in C57BL/6J mice

“…The BNST specifically also undergoes developmental maturation during this early postnatal period (del Abril, Segovia, & Guillamón, 1987;Carter, 2017;Chung, Swaab, & De Vries, 2000;Verney, Gaspar, Febvret, & Berger, 1988), rendering it vulnerable as well to dysfunction instigated by maternal separation stress. In fact, repeated maternal separation during the mouse developmental critical period (i.e., has been shown to alter c-Fos activity patterns in the BNST (Horii-Hayashi et al, 2013), and studies in rats have shown that ELS alters postnatal circuit assembly in the BNST (Card et al, 2005).…”

Novel Rbfox2 isoforms associated with alternative exon usage in rat cortex and suprachiasmatic nucleus

“…Sections containing the SCN were selected, and protein expression was analysed using fluorescence immunohistochemistry 55 . The primary antibodies used for immunohistochemistry (RBFOX2, RBM9 IHC-00199, Bethyl Laboratories Inc., Montgomery, TX, USA; NeuN/RBFOX3, MAB377, Millipore, Temecula, CA, USA), and MeCP2 (4B4, Active Motif, Carlsbad, CA, USA), have been used in our previous studies 19 , 55 , 56 , and have been validated both positively (excitation light-specific detection), and negatively (absence of detection when the primary antibody is omitted). The epitope recognized by IHC-00199 maps to a region in the N-terminal domain (residues 1–50 of hRBFOX2; NP_055124.1) that is 100% conserved between human and rat.…”

Confirmation of the presence of serotonergic cells in the fetal cerebral cortex in cultures and <i>in situ</i>

“…15,16 There are other types of early-appearing neurons in the growing cerebral cortex, such as Cajal-Retzius cells in the marginal zone, which are related to cell migration and to the inside-out cell development pattern of the cortex, 17,18 as well as to GABAergic and tyrosine hydroxylase (TH)-immunopositive neurons in the rat dorsolateral and medial cortex pre-and postnatally. 19,20 It is probable that their phenotypical expression is epigenetically regulated, as in the peripheral nervous system. 21,22 Therefore, cortical neurons may transiently express some enzyme molecules such as TH 20 and other neurotransmitters.…”

“…19,20 It is probable that their phenotypical expression is epigenetically regulated, as in the peripheral nervous system. 21,22 Therefore, cortical neurons may transiently express some enzyme molecules such as TH 20 and other neurotransmitters. 23 Early appearance (gestation days 10 and 11) of GABAergic and noradrenergic neurons is also observed during the pre-and postnatal periods.…”

“…Their expression in the maturing brain may be transient and play an important role in cortical cell differentiation. 20 According to the above-mentioned results, the present work was carried out with the purpose to obtain further information on the presence of serotonergic cells and on the expression of tryptophan-5-hydroxylase (TPH), SERT, 5-HT, the 5-HT 1A receptor, and Pet-1 in the developing cerebral cortex.…”