Anthony LaMantia - US grants

DESCRIPTION (provided by applicant): Work from my laboratory over the previous funding period of this project has shown that mesenchymal/epithelial (M/E) induction, mediated by local signaling molecules including retinoic acid (RA), fibroblast growth factor 8 (FGF8), sonic hedgehog (Shh) and bone morphogenetic proteins (BMPs) is essential for morphogenesis and differentiation of the olfactory pathway during early development of the mammalian forebrain. Our observations raise an essential question: how does induction, mediated by specific molecular signals, influence the identity of specific cell classes in the developing olfactory pathway? To address this question we will evaluate the hypothesis that inductive signaling molecules, acting in the context of M/E interactions, regulate the generation and differentiation of olfactory pathway neurons via their action on molecularly distinct precursor cell populations. We will evaluate this hypothesis in two Specific Aims: the first includes experiments that assess the mechanisms of M/E inductive signaling for establishing precursor populations that give rise to olfactory receptor neurons (ORNs) in the olfactory epithelium, and the second addresses the role of M/E induction in establishing precursors of olfactory bulb interneurons (OBIs: including olfactory granule cells and periglomerular cells). We have developed several in vitro assays, complemented by in vivo approaches, to pursue these aims. Our experiments permit us to manipulate signaling via RA, FGF8, Shh and BMPs using either pharmacological approaches or genetic mutations that result in either loss or gain of function for each signal. Using these tools, we will assess the relationship between signaling via RA, FGF8 and BMP4 and the establishment of molecularly distinct ORN precursor populations as well as the acquisition of functional properties that characterize the mature ORN. In addition, we will evaluate the role of signaling via RA, FGF8, Shh and BMP4 in the context of M/E interaction for OBI development. We will examine the role of signaling patterning the expression of transcriptional regulators associated with OBI precursors in the lateral ganglionic eminence (LGE) as well as facilitating the specific migration of OBI precursors to the rudimentary olfactory bulb as well as their initial differentiation. The results of our experiments will permit us to define the specific contributions of several essential molecular signals to establishing two cell types that must not only be generated during early forebrain development, but that will continue to be generated and integrated into functional circuits from precursor populations that remain in the adult forebrain. Thus, our results will indicate how inductive signals act to define neuronal classes during initial development of a major functional division of the forebrain, the olfactory pathway, as well as how specific signals might contribute to the ongoing regeneration and repair of these forebrain neurons and circuits throughout life.

The ability to control gene expression is essential for neurons of the central nervous system to respond appropriately to changes in the environment. These changes include normal experience as well as compensatory or regenerative changes following injury. In spinal cord pathways that relay and process information about pain, some neurons modify their cellular or functional characteristics in response to experience, painful stimuli or injury. The PI wishes to determine whether the control of gene expression via retinoic acid (vitamin A), a steroid-like hormone that influences the initial development of neurons in dorsal horn of the embryonic spinal cord, contributes to this capacity for change in the mature cord. Preliminary observations show that a distinct subpopulation of cells in lamina I, II and III of the dorsal horn activate gene expression via retinoic acid signaling throughout life. The P.I. proposes to establish the relationship between these retinoid-activated cells and neurons that process and relay pain information in the dorsal spinal cord. Furthermore, the P.I. will evaluate whether retinoic acid activation of gene expression can contribute to changes in transcription in dorsal horn neurons that are seen following injury. The control of gene expression by retinoic acid may be related to the modulation of expression of immediate early genes like c-fos and the subsequent increase in expression of opiod and other peptide neurotransmitters as well as their receptors in the dorsal horn. Accordingly, Dr. Lamantia will determine whether the cells that respond transcriptionally to retinoids have distinct connectional and biochemical properties as well as the capacity to modify gene expression in response to peripheral trauma or deafferentation. Thus, the experiments in this proposal will allow the PI to evaluate critically the hypothesis that a distinctive transcriptional control mechanism used during development of the spinal cord-retinoid mediated transcriptional activation- subsequently contributes to cellular and functional plasticity in specific populations of neurons in the mature spinal cord.

DESCRIPTION (provided by applicant): The 22q11 Chromosomal Microdeletion Syndrome (22q11DS) is the most common survivable deletion syndrome known. In addition to heart, face and limb malformations, 22q11DS is defined by behavioral anomalies and increased susceptibility to several psychiatric diseases including schizophrenia. Apparently, 22q11 haploinsufficiency alters brain development or function, particularly forebrain regions and circuits compromised in behavioral and psychiatric disorders. Nevertheless, the effects of haploinsufficiency on expression or activity of 22q11 genes in the forebrain remain unknown. Our preliminary evidence suggests that 18 of 29 routine 22q11 homologues are expressed throughout the forebrain from early development through adulthood. Accordingly, we will evaluate the hypothesis that there is concerted expression of multiple 22q11 genes in specific cell classes distributed broadly in all forebrain subdivisions. If this is the case, multigenic haploinsufficiency at 22q11, rather than the function of any single gene, might establish a threshold for altered development of specific cell classes or circuits throughout the forebrain. This primary effect may be further amplified by alterations in mesenchymal/epithelial induction that mediates initial differentiation in the forebrain as well as the heart, limbs and face. Moreover, continued expression of 22q11 homologues, perhaps in specific differentiated cell classes, suggests that haploinsufficiency might further compromise function or maintenance of mature forebrain cells and circuit elements. To test our hypothesis we will determine cell-class specificity of a subset of the 22q11 homologues. We selected these genes based upon their significant and sustained expression levels in the developing and adult forebrain, genomic organization, inferred neural function and association with behavioral or psychiatric disorders. In addition, we will analyze the role of mesenchymal/epithelial interactions in modulating 22q11 gene expression. Finally, we will evaluate the consequences of 22q11 haploinsufficiency for expression patterns, levels and cellular phenotype in the developing and adult forebraIn. Thus, our experiments will establish the cellular substrates for the behavioral and psychiatric phenotypes seen in 22q11DS, and give insight into the developmental and genomic mechanisms by which haploinsufficiency at 22q11 compromises forebrain structure and function.

DiGeorge's syndrome; behavioral /social science research tag; laboratory mouse

The ability to rapidly identify and clone genes of interest, synthesize cRNA probes, and resolve regional and cellular expression has facilitated current understanding of cellular diversity in the brain, as well as appreciation of relationships between mutations, gene expression, and aberrant phenotypes. In parallel, quantitative PCR has emerged as a complimentary method to assess variation in levels as well as patterns of expression. These two methods provide a necessary transition from molecular discovery-based approaches such as expression microarray to hypothesis-driven experiments to understand gene function in the brain. The integration of expression analysis with other Core services described in this proposal facilitates the most difficult step in molecular and genetic analyses of CMS development or function: translation of molecular data into cell biological and systemic insight. The expansion of Core 2 services to include quantitative PCR as well as a well-developed library of validated probe templates will help insure the high level of productivity maintained by NINDS-supported investigators who accessed and used core services in the past 4 years.