Central Nervous System Diseases; Mice, Inbred Strains; Mice, Transgenic; Neuroanatomy; Neurosciences; Patients; Regeneration; Developmental Biology; Living Donors; Psychiatry and Psychology
Stem Cell Center, Yale: Stem Cell Genetics
WHRY Pilot Project Program Investigators
Neural stem and progenitor cells give rise to neurons and glia (the central nervous system supporting cells) at both prenatal and postnatal stages of development. The proliferation, migration, and differentiation of neural stem cells into a large variety of neuronal and glial cell types are regulated by a complex array of factors. We study how cell-to-cell contacts and signaling systems govern the behavior of neural stem cells and brain plasticity in the embryonic and postnatal periods. Our goal is to understand whether abnormal proliferation and differentiation of neural stem cells contributes to the genesis of disorders such as autism, Tourette syndrome, and developmental disabilities. We recently founded the The Program in Neurodevelopment and Regeneration. The objectives of this new interdepartmental program are to use induced pluripotent cells (iPSC) as a tool to understand neuronal development in individuals with specific neuropsychiatric disorders. Neuronal development will be recapitulated in vitro by differentiating neuronal progenitors of various CNS lineages from iPSC lines. This will allow us to perform cellular, molecular, genetic, epigenetic and functional studies of these cell lines.
Extensive Research Description
The Vaccarino laboratory has elucidated crucial mechanisms that regulate neural stem cell self-renewal, survival and differentiation. The lab has been redefining the roles of astroglial cells—extremely diverse cellular elements that evolve from a primary role of neural progenitors during embryonic development, to essential partners in neuronal migration, maturation, axon guidance. These cells have essential metabolic function in the adult brain and retain ability to divide and act as precursors.
In the 1990’s Vaccarino and colleagues reported that an extracellular protein called Basic Fibroblast Growth Factor 2 (FGF2) increases the number of progenitors for excitatory cortical neurons in vitro (Vaccarino et al., 1995) and that a single microinjection of FGF2 into the cerebral ventricles of rat embryos doubled the number of excitatory neurons generated during cortical development (Vaccarino et al., 1999). This was the first evidence that a single factor can elicit a permanent increase in cerebral cortical size and neuron number in a mammalian species. Conversely, using knockout mice, Vaccarino and colleagues demonstrated that the Fgf2 gene product is essential for the generation of a normal number of excitatory neurons in the cerebral cortex by increasing cell proliferation in the cortical neuroepithelium (Vaccarino et al., 1999; Raballo et al., 2000; Korada et al., 2002). Subsequent work of demonstrated that FGF receptors are essential for normal telencephalic development. The disruption of the Fgfr1 gene is sufficient to thwart cell proliferation within the hippocampal primordium, causing lifelong hippocampal atrophy (Ohkubo et al., 2004). In contrast, the antagonism of all FGF receptors is required to disrupt cortical development. Driving a dominant negative Fgfr1 to the cortical primordium resulted in decreased generation and lower numbers of glutamatergic pyramidal neurons, which form the primary scaffold of the cortex, and cortical atrophy (Shin et al., 2004).
In the evolution of this research, Vaccarino and her colleagues discovered that FGF receptors play an indirect but crucial role in the development of inhibitory neurons. A lower number of inhibitory neurons containing parvalbumin was observed in the adult cerebral cortex of both Fgfr1 and Fgfr2 null mutant mice (Muller Smith et al., 2008). Recent data suggest that the GABA neuron defect is attributable to their deficient maturation or survival during early postnatal development. Further, the stereotypic locomotor hyperactivity exhibited by FGF receptor mutant mice was inversely proportional to the number of parvalbumin inhibitory neurons in their cerebral cortex (Muller Smith et al., 2008). Interestingly, parallel work in postmortem human brains performed by the same lab showed that the same parvalbumin inhibitory neurons are decreased in number within the caudate nucleus of patients with Tourette syndrome (Kalanithi et al., 2005). It is also known that cortical parvalbumin neurons are decreased in schizophrenia and other psychotic disorders. The lab is now focused on elucidating the molecular and cellular mechanisms that control parvalbumin neuron differentiation and survival throughout early postnatal development. These processes may require paracrine astroglia-neuron signaling, which requires FGF receptors.
Another idea that has been put forward by the Vaccarino laboratory is that cortical neurogenesis does not abruptly stop at birth but that the immature brain has the capacity to produce new cortical neurons, a process that is enhanced by injury. The lab has observed that after neonatal hypoxia, the excitatory neuron losses spontaneously recover in the cerebral cortex (Fagel et al., 2006, and Fagel et al., 2009). During this phase there is a prominent increase in cell proliferation and neurogenesis, along with an increase in expression of FGF2 and FGFR1 in the forebrain germinal zones (Ganat et al, 2002). The proliferation, neurogenesis and ensuing neuronal recovery are both nearly absent in Fgfr1 conditional null mutant mice. This represents the first evidence that excitatory neurons can recover after a postnatal insult and that FGFR1 in astroglial cells is centrally involved in this process (Fagel et al, 2009).
To further study the role of glial stem/progenitor cells in regenerative processes, the lab has generated the GFAP-CreERT2 transgenic line, in which the Cre recombinase can be transiently induced by a drug injection in astroglial cells (Ganat et al, 2006). This mouse model allows investigators to identify and follow astroglial cells and their progeny. These cells can be also genetically altered in a temporally controlled manner. In recognition of this work, the laboratory was recently awarded a grant from the State of Connecticut to study the gene expression and fate potential of astroglial cells after hypoxic insult.
Studies are underway to reveal the mechanisms and pathways that are involved in brain adaptation and self-repair after this insult. Parallel experiments will elucidate astroglia-specific transcripts that allow these cells to undergo dramatic fate transition from quiescent to reactive/proliferative, and from glial to neuron.
The Vaccarino research has contributed key insights into the processes that regulate the number and differentiation of excitatory and inhibitory neurons in both prenatal and postnatal development. They have proposed that the balance between excitatory and inhibitory neurons in the cerebral cortex and connected regions is regulated by developmental genes (i.e., Fgf) as well as by environmental factors (i.e., hypoxia) acting at crucial stages in development. Disruption of this delicate balance produces predictable alterations in psychomotor behavior in our animal models. These animal models mimic aspects of neuropsychiatric disorders (i.e., hyperactivity, impulsivity, poor learning) and can be used to study ways to repair or reverse these deficits. The cerebral cortex and connected regions engage patterns of goal-directed behavior that are assembled during infancy. Concomitantly, the cortex represses impulsive, unwanted behaviors. Poor organization and control of cognitive and motor processes due to disrupted cortical development may be shared in various ways by schizophrenia, autism, ADHD and Tourette’s Syndrome. Dr. Vaccarino is the Director of the Developmental Neurobiology Laboratory (Vaccarino Lab) at the Child Study Center, Yale University School of Medicine. She and a group of collaborating investigators recently formed the interdepartmental Program in Neurodevelopment and Regeneration.
In the Developmental Neurobiology Laboratory the Vaccarino team focuses on two main areas:
1. Proliferation and differentiation of neural stem cells (NSCs) during prenatal and postnatal stages of cerebral development and in response to injury.
a. Role of Fibroblast Growth Factors (FGFs) in NSC differentiation. Using gain or loss of function for FGFs and their receptors in NSC, we have shown that prenatal loss of FGF/FGFRs causes altered brain size and imbalances in excitatory and inhibitory neuron types, with long-term repercussions in the animal’s behavior. In addition, FGFs are required postnatally for brain regeneration and recovery after perinatal hypoxic injury.
b. Diversity and function of astroglia. Astroglial cells comprise a diversity of cellular phenotypes, from radial glial progenitors in prenatal development to astrocytes in adult brains. These cells undergo plastic changes during development and aging; postnatal NSCs are modified astroglial cells. We are analyzing the transcriptome of astroglial cells in normal development and after hypoxia or environmental perturbations. The function of specific gene products in astroglia is investigated by inducible cre recombination, both at the cellular and organism level.
2. Cellular alterations in neuropsychiatric disorders.
a. Mouse models. The effects of perinatal hypoxia on brain development. We study an animal model of perinatal hypoxia, which models cognitive handicap common in premature children. We have shown that mice raised in hypoxic conditions have a persistent deficit in inhibitory interneurons in the cerebral cortex. We are investigating the cellular and molecular mechanisms that mediate brain injury and brain repair, focusing on the role of astroglial cells in orchestrating recovery.
b. Postmortem analyses of the human brain. An excitatory/inhibitory neuronal imbalance in specific forebrain systems due to disparate etiologies can be the common pathogenesis of childhood neuropsychiatric disorders such as Tourette’s syndrome and autism. For example, we have shown that individuals with Tourette’s syndrome (TS) have losses of Parvalbumin and cholinergic interneurons in specific regions of the striatum. The hypothesis is that altered postnatal maturation/survival of subsets of inhibitory neurons in cortico-basal ganglia circuits causes specific alterations in synchronous neuronal firing that may model disorders of the TS spectrum.
c. Induced pluripotent stem cells (iPSC). To recapitulate the dynamics of human neuronal development in an in vitro system, we will generate induced pluripotent cells (iPSC) by “reprogramming” somatic cells into a pluripotent state. These cells can be induced to differentiate into neural progenitors and then into forebrain neurons and glia. Using somatic cells from patients with neuropsychiatric disorders will allow us to investigate disease and patient-specific features of the neural differentiation process.
The Program in Neurodevelopment and Regeneration involves faculty from Child Study Center, Genetics, Neurobiology, Psychiatry, and Pediatrics.The objectives of this new interdepartmental program are to use induced pluripotent cells (iPSC) as a tool to understand neuronal development in individuals with specific neuropsychiatric disorders.
Neuronal development will be recapitulated in vitro by differentiating neuronal progenitors of various CNS lineages from iPSC lines. This will allow different investigators participating in this program to perform cellular, molecular, genetic, epigenetic and functional studies of these cell lines.
The scientific goals of the Program are:
1. A detailed cellular and functional study of the particular features of neurons and glia derived from patient-specific iPSC lines to understand the biological mechanisms of neuropsychiatric disorders.
2. Correlate cellular and functional events in neural development with underlying changes in genomic sequence, epigenomic imprinting and regulation of gene expression.
- Hypoxia-induced developmental delays of inhibitory interneurons are reversed by environmental enrichment in the postnatal mouse forebrain. M Komitova, D Xenos, N Salmaso, KM Tran, T Brand, ML Schwartz, L Ment, FM Vaccarino (2013). J Neurosci 33: 13375-87.
- Functional genomic screen of human stem cell differentiation reveals pathways involved in neurodevelopment and neurodegeneration. Y Zhang, VP Schulz, BD Reed, Z Wang, X Pan, J Mariani, G Euskirchen, MP Snyder, FM Vaccarino, N Ivanova, SM Weissman, AM Szek
- Cortical gyrification induced by fibroblast growth factor 2 in the mouse brain. BG Rash, S Tomasi, HD Lim, CY Suh, FM Vaccarino (2013). J Neurosci 33: 10802-14.
- Prenatal stress delays inhibitory neuron progenitor migration in the developing neocortex. HE Stevens, T Su, Y Yanagawa, FM Vaccarino (2013). Psychoneuroendocrinology 38: 509-21.
- Age-related changes of gene expression in the neocortex: preliminary data on RNA-Seq of the transcriptome in three functionally distinct cortical areas. OY Naumova, D Palejev, NV Vlasova, M Lee, SY Rychkov, ON Babich, F M Vaccarino, EL Grigorenko (2012).
- Abyzov, A., Mariani, J., Palejev, D., Zhang, Y., Haney, M.S., Tomasini, L., Ferrandino, A., Rosenberg Belmaker, L., Szekely, A., Wilson, M., Kocabas, A., Calixto, N.E., Grigorenko, E.L., Huttner, A., Chawarska, K., Weissman, S., Eckehart Urban, A., Gerste
- Mariani, J., Simonini, M.V., Palejev, D., Tomasini, L., Coppola, G., Szekely, A., Horvath, T., & Vaccarino, F.M. (2012). Modeling human cortical development in vitro using induced pluripotent stem cells. Proceedings of the National Academy for Sciences U
- Stevens, H.E., Mariani, J., Coppola, G., & Vaccarino, F.M. (in press). Neurobiology meets genomic science: The promise of human induced pluripotent stem cells. In Grigorenko, E. L., & Cicchetti, D. (Eds.), The contribution of genetic/genomic sciences to d
- Rash, B.G., Lim, H.D., Breunig, J.J., and Vaccarino, F.M.: FGF signaling expands embryonic cortical surface area by regulating Notch-dependent neurogenesis. J Neurosci, 31: 15604-17, 2011.
- Vaccarino, F.M., Urban, A.E., Stevens, H.E., Szekely, A., Abyzov, A.,Grigorenko, E.L., Gerstein, M.,Weissman, S.: Annual Research Review: The promise of stem cell research for neuropsychiatric disorders. J Child Psychol Psychiatry, Ed. 2011/01/06, Jan 4.
- Silbereis, J., Heintz, T., Taylor, M.M., Ganat, Y.M., Ment, L.M., Bordey, A., Vaccarino, F.M.: Astroglial cells in the external granular layer are precursors of cerebellar granule neurons in neonates. Molecular Cell. Neurosci. (2010) 44:362-373.
- Kataoka, Y., Kalanithi, P.S.A., Grantz, H., Schwartz, M.L., Saper, C., Leckman, J.F., Vaccarino, F.M.: Decreased Number of Parvalbumin and Cholinergic Interneurons in the Striatum of Individuals with Tourette Syndrome. J. Comp. Neurol. (2010) 518:277-291
- Stevens, H. E., Smith, K. M. , Maragnoli, M. E. , Fagel, D., Borok, E., Shanabrough, M., Horvath, T. L., Vaccarino, F. M.: Fgfr2 is required for the development of the medial prefrontal cortex and its connections with limbic circuits. J. Neurosci. (2010)
- Vaccarino, F. M., Smith, K.M.: Increased Brain Size in Autism-What it Will Take to Solve a Mystery. Biological Psychiatry 66:313-315, 2009. http://www.ncbi.nlm.nih.gov/pubmed/19643218?dopt=Citation
- Vaccarino, F. M., Smith, K.M., Stevens, H.: Regulation of cerebral cortical size and neuron number by Fibroblast Growth Factors: Implication for autism. Journal of Autism and Developmental Disorders, 39:511-20,2009. http://www.springerlink.com/content/ym
- Silbereis, J, Cheng, E, Ganat, YM, Ment, LM, Vaccarino, FM: Precursors with GFAP promoter activity transiently generate GABA interneurons in the postnatal cerebellum. Stem Cells 27:1152-1163, 2009. http://www.ncbi.nlm.nih.gov/pubmed/19418461?dopt=Citation
- Fagel, D.M., Silbereis, J., Ohkubo, Y., Ganat, Y., Ment, L.R., Vaccarino, F.M.: Fgfr-1 Is Required For Cortical Regeneration and Repair following Perinatal Hypoxia. Journal of Neuroscience (2009) 29(4):1202-1211. http://www.ncbi.nlm.nih.gov/pubmed/19176
- Smith, KM, Fagel, DM, Stevens, HE, Rabenstein, RL, Maragnoli, ME, Ohkubo, Y, Picciotto, MR, Schwartz, ML, Vaccarino, FM: Deficiency in inhibitory cortical interneurons associates with hyperactivity in Fgfr1 mutant mice. Biological Psychiatry, 2008, 63:95
- Ganat, Y.M., et al. (2006). Early postnatal astroglial cells produce multilineage precursors and neural stem cells in vivo. J. Neurosci. 26:8609-8621. http://www.ncbi.nlm.nih.gov/pubmed/16914687?dopt=Citation
- Fagel DM, Ganat Y, Silbereis J, Ebbitt T, Stewart W, Zhang H, et al. Cortical neurogenesis enhanced by chronic perinatal hypoxia. Exp Neurol 2006199(1):77-91. http://www.ncbi.nlm.nih.gov/pubmed/15916762?dopt=Citation
- Smith, K.M., et al. (2006). Midline radial glia translocation and corpus callosum formation require FGF signaling. Nat. Neurosci. 9:787-797. http://www.ncbi.nlm.nih.gov/pubmed/16715082?dopt=Citation
- Kalanithi PS, Zheng W, Kataoka Y, DiFiglia M, Grantz H, Saper CB, et al. Altered parvalbumin-positive neuron distribution in basal ganglia of individuals with Tourette syndrome. Proceedings of the National Academy of Sciences U S A 2005;102(37):13307-12.
- Ohkubo, Y, Uchida, AO, Shin, D, Partanen, J. and Vaccarino, FM: Fibroblast Growth Factor Receptor 1 is required for the proliferation of hippocampal progenitor cells and for hippocampal growth in mouse. J Neurosci, 24: 6057-6069, 2004. http://www.ncbi.nlm
- Zheng, W, Nowakowski, R, and Vaccarino, FM: Fgf2 is required for maintaining the neural stem cell pool in the mouse brain subventricular zone. Developmental Neurosci, 26:181-196, 2004.
- Shin, D, Korada, S, Raballo, R, Shashikant, CS, Simeone, A, Taylor, J, and Vaccarino, FM: Loss of Glutamatergic pyramidal neurons in frontal and temporal cortex due to attenuation of FGFR1 signaling is associated with spontaneous hyperactivity in mice. J
- Ganat, Y, Soni S, Chacon, M, Schwartz, ML and Vaccarino, FM: Chronic hypoxia upregulates Fibroblast Growth Factor (Fgf) ligands in the perinatal brain and induces Fgf-responsive radial glial cells in the subependymal zone. Neuroscience,112:977-991, 2002.
- Korada, S, Zheng, W, Basilico, C, Schwartz, ML and Vaccarino, FM: Fgf2 is necessary for the growth of glutamate projection neurons in the anterior neocortex. J Neurosci, 22:863-875, 2002.
- Raballo, R, Rhee, J, Lyn-Cook, R, Leckman, JF, Schwartz, M.,Vaccarino, FM: Basic Fibroblast Growth Factor (Fgf2) is necessary for cell proliferation and neurogenesis in the developing cerebral cortex J Neurosci, 20:5012-5023, 2000. http://www.ncbi.nlm.ni
- Vaccarino, FM, Schwartz, ML, Raballo, R, Nilsen, J, Rhee, J, Zhou, M, Doetschman, T, Coffin, JD, Wyland, J, Hung, Y-T: Changes in cerebral cortex size are governed by Fibroblast Growth Factor during embryogenesis. Nature Neurosci, 3: 246-253, 1999. http:/