Distinct types of relay neurons in the hindbrain process somatosensory or viscerosensory information. How neurons choose between these two fates is unclear. We show here that the homeobox gene Lbx1 is essential for imposing a somatosensory fate on relay neurons in the hindbrain. In Lbx1 mutant mice, viscerosensory relay neurons are specified at the expense of somatosensory relay neurons. Thus Lbx1 expression distinguishes between the somatosensory or viscerosensory fate of relay neurons.

The Olig3 gene encodes a bHLH factor that is expressed in the ventricular zone of the dorsal alar plate of the hindbrain. We found that the Olig3 + progenitor domain encompassed subdomains that co-expressed Math1, Ngn1, Mash1 and Ptf1a. Olig3 + cells give rise to neuronal types in the dorsal alar plate that we denote as class A neurons. We used genetic lineage tracing to demonstrate that class A neurons contribute to the nucleus of the solitary tract and to precerebellar nuclei. The fate of class A neurons was not correctly determined in Olig3 mutant mice. As a consequence, the nucleus of the solitary tract did not form, and precerebellar nuclei, such as the inferior olivary nucleus, were absent or small. At the expense of class A neurons, ectopic Lbx1 + neurons appeared in the alar plate in Olig3 mutant mice. By contrast, electroporation of an Olig3 expression vector in the chick hindbrain suppressed the emergence of Lbx1 + neurons. Climbing fiber neurons of the inferior olivary nucleus express Foxd3 and require Olig3 as well as Ptf1a for the determination of their fate. We observed that electroporation of Olig3 and Ptf1a expression vectors, but not either alone, induced Foxd3. We therefore propose that Olig3 can cooperate with Ptf1a to determine the fate of climbing fiber neurons of the inferior olivary nucleus.

. Osmolality and solute composition are strong regulators of AQP2 expression in renal principal cells. Am J Physiol Renal Physiol 284: F189-F198, 2003. First published August 13, 2002; 10.1152/ajprenal.00245.2002The water permeability of the renal collecting duct is regulated by the insertion of aquaporin-2 (AQP2) into the apical plasma membrane of epithelial (principal) cells. Using primary cultured epithelial cells from the inner medulla of rat kidney (IMCD cells), we show that osmolality and solute composition are potent regulators of AQP2 mRNA and protein synthesis, as well as the classical cAMP-dependent pathway, but do not affect the arginine vasopressin-induced AQP2 shuttle. In the presence of the cAMP analog dibutyryl cAMP (DBcAMP, 500 M), NaCl and sorbitol, but not urea, evoked a robust increase of AQP2 expression in IMCD cells, with NaCl being far more potent than sorbitol. cAMP-responsive elementbinding protein phosphorylation increased with DBcAMP concentrations but was not altered by changes in osmolality. In the rat and human AQP2 promoter, we identified a putative tonicity-responsive element. We conclude that, in addition to the arginine vasopressin/cAMP-signaling cascade, a further pathway activated by elevated effective osmolality (tonicity) is crucial for the expression of AQP2 in IMCD cells, and we suggest that the effect is mediated via the tonicityresponsive element. osmolality; aquaporin-2; kidney; gene regulation; toxicity responsive enhancer THE WATER CHANNEL aquaporin-2 (AQP2), expressed in epithelial (principal) cells of renal collecting ducts, is required for the vasopressin-dependent concentration of urine (7). AQP2 abundance increases from the kidney cortex to the inner region of the kidney medulla (29), as does osmolality. The water permeability of the inner medullary collecting duct (IMCD) is rapidly regulated (within minutes) by the antidiuretic hormone arginine vasopressin (AVP), which binds to heptahelical vasopressin V 2 receptors (V 2 R), located mainly in the basolateral plasma membrane of principal cells. Activation of the V 2 R causes stimulation of adenylyl cyclase via the G protein G S , leading to elevation of cAMP. The subsequent activation of protein kinase A (PKA) initiates the translocation of AQP2-bearing vesicles from the cytosol to the plasma membrane, in which AQP2 is inserted by an exocytosis-like process (short-term regulation) (D. Lorenz, A. Krylov, V. Hagen, J. Zipper, W. Rosenthal, P. Pohl, and K. Maric, unpublished observations; 30). In addition, the signaling cascade described above governs the expression of AQP2 (long-term regulation) by PKA-mediated phosphorylation of the transcription factor cAMP-responsive element (CRE)-binding protein (CREB) (13,16). Given the fact that AQP2 biosynthesis is usually shut off shortly after IMCD cells are established in primary culture (10), most studies on AQP2 long-term regulation have been performed using animal models (29, 37). We recently established primary cultured IMCD cells as a model system (17,18,21,22). Thes...

Monoaminergic neurons include the physiologically important central serotonergic and noradrenergic subtypes. Here, we identify the zinc-finger transcription factor, Insm1, as a crucial mediator of the differentiation of both subtypes, and in particular the acquisition of their neurotransmitter phenotype. Insm1 is expressed in hindbrain progenitors of monoaminergic neurons as they exit the cell cycle, in a pattern that partially overlaps with the expression of the proneural factor Ascl1. Consistent with this, a conserved cis-regulatory sequence associated with Insm1 is bound by Ascl1 in the hindbrain, and Ascl1 is essential for the expression of Insm1 in the ventral hindbrain. In Insm1-null mutant mice, the expression of the serotonergic fate determinants Pet1, Lmx1b and Gata2 is markedly downregulated. Nevertheless, serotonergic precursors begin to differentiate in Insm1 mutants, but fail to produce serotonin because of a failure to activate expression of tryptophan hydroxylase 2 (Tph2), the key enzyme of serotonin biosynthesis. We find that both Insm1 and Ascl1 coordinately specify Tph2 expression. In brainstem noradrenergic centres of Insm1 mutants, expression of tyrosine hydroxylase is delayed in the locus coeruleus and is markedly deficient in the medullary noradrenergic nuclei. However, Insm1 is dispensable for the expression of a second key noradrenergic biosynthetic enzyme, dopamine β-hydroxylase, which is instead regulated by Ascl1. Thus, Insm1 regulates the synthesis of distinct monoaminergic neurotransmitters by acting combinatorially with, or independently of, Ascl1 in specific monoaminergic populations.

The homeodomain factor Lbx1 is expressed in postmitotic neurons in the ventral alar plate of the developing hindbrain extending from rhombomere 2 to the caudal medulla.It has been shown that Lbx1 distinguishes two major programs of neuronal differentiation in the dorsal spinal cord (class A, Lbx− and class B, Lbx+).Here, we introduce a classification of the neuronal subtypes emerging in the alar plate of the hindbrain according to their transcription factor profiles. Lineage tracing allowed us to define derivatives of Lbx+ neurons, which include the spinal trigeminal nucleus and GABAergic neurons in the nucleus of solitary tract. The nucleus of the solitary tract is a major brainstem relay station for visceral sensory input and modulates the frequency of breathing and cardiac contraction, whereas the spinal trigeminal nucleus receives somatosensory input. The loss of Lbx1 leads to an abnormal molecular identity and to a mis-specification of neurons participating in the formation of these sensory relay centers. In Lbx1 mutant mice the spinal trigeminal nucleus is greatly reduced or lost and the nucleus of solitary tract is enlarged and exhibits an abberant neuronal circuitry. This is accompanied by long periods of apnea and a reduced frequency of cardiac contraction. Lbx1 is therefore an important determinant in the specification of hindbrain neurons that process sensory information.In mammals, VIP is expressed during morphogenesis of nervous and genital systems, suggesting a trophic action and neuronal survival. In birds, the intercellular signals of VIP are regulated via G-protein-coupled receptors, in neural and reproductive tissues. In mammalian two VIPR subtypes have identified; in avian species VIP type I was mainly expressed.The aim of this study was to identify the distribution of VIP and its receptor during morphogenesis in chicken ovary by immunolocalization techniques. Left ovary from embryos and post-hatching birds were used.VIP immunoreactivity was observed in cortex since 13th incubation day, inside of celomic epithelium. Later VIP was detected in pregranulosa cells around the oocytes; in medulla over interstitial steroidogenic cumuli, lacunary channels and somas and nerve fibers. In post-hatching animals, distribution of VIP originated a gradient from cortex to medulla. VIP was detected in granulosa sheet and some oocytes in immature follicles.In early stages of embryogenesis, VIPR was observed in cortex, on some epithelial cells, pregranulosa cells and oocytes cumulus and on steroidogenic cells islets in ovarian medulla. In post-hatching animals, 1st, 2nd, 4th and 7th post-hatching day, VIPR immunostaining was detected in oocytes and granulosa cells; neuronal somas and fibers.This study suggests a functional effect of VIP and VIPR on gonadal development. Acknowledgement

(ER) and the Golgi apparatus. Altered protein glycosylation can manifest in serious, sometimes fatal malfunctions. We recently showed that mutations in GDP-mannose pyrophosphorylase A (GMPPA) can cause a syndrome characterized by alacrima, achalasia, mental retardation, and myopathic alterations (AAMR syndrome). GMPPA acts as a feedback inhibitor of GDP-mannose pyrophosphorylase B (GMPPB), which provides GDP-mannose as a substrate for protein glycosylation. Loss of GMPPA thus enhances the incorporation of mannose into glycochains of various proteins, including α-dystroglycan (α-DG), a protein that links the extracellular matrix with the cytoskeleton. Here, we further characterized the consequences of loss of GMPPA for the secretory pathway. This includes a fragmentation of the Golgi apparatus, which comes along with a regulation of the abundance of several ER- and Golgi-resident proteins. We further show that the activity of the Golgi-associated endoprotease furin is reduced. Moreover, the fraction of α-DG, which is retained in the ER, is increased. Notably, WT cells cultured at a high mannose concentration display similar changes with increased retention of α-DG, altered structure of the Golgi apparatus, and a decrease in furin activity. In summary, our data underline the importance of a balanced mannose homeostasis for the secretory pathway.

Vasopressin (AVP) induces antidiuresis via its renal V2 receptors but also acts as a neuropeptide through activation of central V1a and V1b receptors (V1aR, V1bR). Despite a large body of functional evidence for roles of these receptors in motivated behavior, little information is available on their distribution in the brain tissue. This study was designed to provide a comparative analysis of the V1aR and V1bR distribution patterns in rodent and human brain tissues using new antisera.Antibodies to individual AVP receptors were generated and validated using respective mouse knockout tissues or overexpression of the human V1aR and V1bR in HEK293 cells. Distribution of the receptors was studied by immunofluorescence in vivo in perfused rat or mouse brains and human brain samples obtained from clinical section. Confocal microscopy or super resolution imagingwere applied for signal detection. AVP receptors expressing cell types were identified by double‐labeling for markers of neurons (CTIP2 or SATB2) or glial cells (GFAP).Application of the anti‐V1aR antibody on rat or mouse brain sections produced signal in pyramidal cells of the isocortical layers II and V, astrocytes, neurons of the medial habenula, the suprachiasmatic and supraoptic hypothalamic nuclei, as well as in the CA2 and CA3 fields of hippocampus, whereas no significant signal was detected in the choroid plexus. V1bR signal was present in pyramidal cells of the isocortical layers II and V, neurons of the medial and lateral habenula, the suprachiasmatic and supraopticus nuclei of hypothalamus and in basolateral cellular aspects of the choroid epithelia. Evaluation of human brain samples revealed similar V1aR and V1bR distribution patterns, as compared to rodents.In summary, our results document the distribution patterns of V1aR and V1bR in the rodent brain, thus corroborating previous functional data on the role of AVP in motivated behavior.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Whole animal behaviors require precise wiring of the nervous system. Yet how neuronal wiring translates into defined behaviors is not well understood. To study the interplay between nervous