2008 — 2012 |
Zakharenko, Stanislav S |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Trans-Synaptic Interactions During Synaptic Plasticity in the Hippocampus @ St. Jude Children's Research Hospital
Project summary: To understand learning and memory, we must first elucidate the molecular machinery that regulates synaptic plasticity in the CNS. Recent results indicate that long-term potentiation (LTP), a major form of synaptic plasticity, is not a unitary process, even at a single synapse. LTP is expressed by independent presynaptic and postsynaptic mechanisms at excitatory synapses between CA3 and CA1 pyramidal neurons in the hippocampus. Given that the induction of plasticity at these synapses involves the influx of calcium ions into postsynaptic CA1 neurons, the expression of the presynaptic component of LTP suggests the existence of a retrograde signal that transfers a signal from the postsynaptic CA1 neuron to the presynaptic CA3 cell. Various molecular mechanisms have been implicated in retrograde signaling during LTP; however, none alone satisfies all of the requirements. In this proposal, we will examine the roles of nitric oxide, integrins, and neuronal adhesion molecule L1 in mechanisms of retrograde signaling during LTP. To accomplish these goals, we will directly visualize functional changes using fluorescent indicators of presynaptic function at the level of a single synapse. Two-photon laser scanning microscopy and 2-photon laser scanning uncaging techniques will be used in combination with electrophysiological tools in acute hippocampal slices from transgenic mice expressing various fluorescent markers in pyramidal neurons. Relevance: Long-term potentiation (LTP) is the use-dependent enhancement of the signal between neurons in the brain. LTP is thought to be a key process that regulates learning and memory. To better understand learning and memory and further the discovery of ways to prevent disease from disrupting these essential abilities in humans, we will investigate the molecular bases of LTP in mice with appropriate genetic mutations.
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1 |
2012 — 2016 |
Zakharenko, Stanislav S |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Identification of Synaptic Mechanisms of 22q11 Deletion Syndrome @ St. Jude Children's Research Hospital
DESCRIPTION (provided by applicant): The 22q11 deletion syndrome (22q11DS), also known as velocardiofacialsyndrome or DiGeorge syndrome, is the most common microdeletion syndrome in humans. Cognitive deficits occur in virtually all patients with 22q11DS, and schizophrenia or schizoaffective disorder develops in approximately 30% during their adolescence or early adulthood. Deficits in learning and memory have been identified in patients with 22q11DS and in Df(16)1/+ mice, the mouse model of this disease. However, the cellular mechanisms and gene(s) responsible for these deficits remain unknown. Recently, we discovered that long-term potentiation (LTP) of synaptic transmission, a major form of synaptic plasticity and cellular substrate of certain forms of learning and memory, is substantially altered in Df(16)1/+ mice. We determined that these changes are caused by the abnormal presynaptic function at excitatory synapses. Further experiments revealed that the increase in presynaptic function was caused by the deletion of 2 genomic regions within the large microdeletion, Df(16)2 and Df(16)5. Screening of mice with deletions of individual genes within the Df(16)2 region revealed that a deletion of the microRNA-processing gene Dgcr8 upregulates sarco(endo)plasmic reticulum ATP-ase 2 (SERCA2) in excitatory neurons and leads to abnormal neurotransmitter release and LTP. The identity of the culprit gene within the Df(16)5 region remains unknown. In this application, we propose to identify the microRNA(s) responsible for the upregulation of SERCA2 and the defects in synaptic plasticity by using electrophysiological and molecular tools, two-photon laser scanning microscopy, and two-photon uncaging. Using knockout mice recently developed in our laboratory, we will also identify the culprit gene(s) within the Df(16)5 region. Finally, we propose to test the role of the endoplasmic reticulum in presynaptic phenotypes of mouse models of 22q11DS. This information will provide a framework for the future development of therapeutic interventions to prevent or alleviate cognitive deficits in patients with 22q11DS. PUBLIC HEALTH RELEVANCE: Heterozygous deletions within the 22q11 chromosome substantially increase an individual's risk for schizophrenia. Cognitive function is characteristically impaired in patients and mouse models that carry these deletions. To better understand these deficits and find the causal genes and downstream signaling pathways, we will investigate the mechanisms of synaptic abnormalities in mutant mice that model 22q11 deletion syndrome.
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1 |
2013 — 2021 |
Zakharenko, Stanislav S |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Synaptic Mechanisms of Auditory Memory @ St. Jude Children's Research Hospital
Abstract: Primary sensory cortices analyze sensory information and store information about learned sensory experiences. The auditory cortex (ACx) acquires and retains memory traces about the behavioral significance of selected sounds. During learning, the tuning properties of ACx neurons undergo activity-dependent changes. This cortical map plasticity, which is believed to be a substrate of auditory memory, is characterized by the facilitation of responses to behaviorally important tones. In juvenile animals, cortical map plasticity in the ACx can be induced by passive environmental enrichment with a certain sound. In rodents, juvenile cortical map plasticity is limited to a few postnatal days (i.e., the early critical period). In mature animals, cortical map plasticity can be induced only if tones are behaviorally important or paired with the activation of modulatory (e.g., cholinergic, dopaminergic, noradrenergic) projections. During the previous funding period, we determined that cortical map plasticity is encoded by the same mechanisms as long-term potentiation (LTP) and long-term depression (LTD) at thalamocortical (TC) excitatory synapses. TC projections are the major sensory input to the neocortex and contribute to the formation of cortical maps. In brain slices, we showed that TC synaptic plasticity is not lost after the early critical period, instead a gating mechanism is acquired that can be released by activating cholinergic receptors on presynaptic terminals. Once gating is released, LTP/LTD at TC synapses and cortical map plasticity in vivo occur in animals aged beyond the early critical period. Adenosine machinery, consisting of adenosine-producing ecto-5'-nucleotidase (Nt5e) and A1 adenosine receptors (A1Rs), provides the gating. Juvenile plasticity can be reestablished in adults, if acoustic stimuli are paired with disruption of Nt5e or A1R signaling in the auditory thalamus. This plasticity occurs in cortical maps and individual ACx neurons of awake adult mice and is associated with long-term improvement in tone-discrimination abilities. In this competitive renewal, we propose to test our hypothesis that the adenosine machinery in the thalamus is the master mediator that transmits information from modulatory projections to the thalamus during ACx map plasticity in adults. In Aim 1, we will induce cortical map plasticity in adults by pairing sounds with activation of modulatory projections while activating or deactivating the gating mechanism. In Aim 2, we will explore the molecular mechanisms of terminating the early critical period by investigating age dependency of adenosine production. In Aim 3, we will determine time scales of the gating mechanisms. Using fast-scan cyclic voltammetry in awake mice, we found that adenosine is transiently released in the auditory thalamus and cortex in response to sound. We propose to elucidate the mechanisms and kinetics of this sound-evoked adenosine release before and after the early critical period and determine how it affects spiking in thalamic relay and cortical neurons during sound stimulation. Knowledge gained from these studies will provide the basis for future elucidation of the cellular and molecular mechanisms of auditory memory.
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1 |
2013 — 2020 |
Zakharenko, Stanislav S |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. R56Activity Code Description: To provide limited interim research support based on the merit of a pending R01 application while applicant gathers additional data to revise a new or competing renewal application. This grant will underwrite highly meritorious applications that if given the opportunity to revise their application could meet IC recommended standards and would be missed opportunities if not funded. Interim funded ends when the applicant succeeds in obtaining an R01 or other competing award built on the R56 grant. These awards are not renewable. |
Towards Understanding Cellular Mechanisms of Positive Symptoms of Schizophrenia @ St. Jude Children's Research Hospital
DESCRIPTION (provided by applicant): Schizophrenia (SCZ) is a devastating disease that affects approximately 1% of the world's population and is characterized by a constellation of symptoms that include hallucinations and delusions (positive symptoms), antisocial behavior and blunted emotions (negative symptoms), deficits in working memory, executive function, and learning and memory (cognitive symptoms). The mechanisms underlying these symptoms remain unknown, mostly due to the lack of valid experimental approaches to model this disease. The 22q11.2-deletion syndrome (22q11DS), also known as velocardiofacialsyndrome or DiGeorge syndrome, is the most common microdeletion syndrome in humans. SCZ arises in approximately 30% of patients with 22q11DS during their adolescence or early adulthood. Mouse models of 22q11DS have been constructed and validated by replicating deficits in working memory, learning and memory, and other symptoms. Using these mutant mice, we and others have identified cellular and molecular mechanisms underlying the cognitive symptoms of 22q11DS. However, self-reported symptoms such as hallucinations cannot be convincingly modeled in mice. In this application, we propose to test the predictions of several recent neuroscience theories and human imaging data that hallucinations result from deficiencies in thalamocortical (TC) pathways that project to the sensory cortices. In our preliminary experiments in brain slices and in vivo, we found that mouse models of 22q11DS have substantial deficits in synaptic transmission and short-term plasticity at TC pathways to the auditory cortex. In this proposal, we will use single-cell electrophysiology, 2-photon imaging, 2-photon glutamate uncaging, optogenetics, and molecular tools to identify the cellular and molecular mechanisms of TC deficiencies in mouse models of 22q11Ds. Using multiple available strains of mutant mice that carry deletions of clusters of genes or individual genes that map within the large 22q11 microdeletion, we will identify the gene(s) whose deletion underlies TC deficits in these mice. We will also perform in vivo 2-photon imaging to observe abnormal spontaneous activity in individual neurons of the auditory cortex. Abnormal neuronal activity in the auditory cortex has been reported in patients who experience auditory hallucinations, which are most predominant in SCZ. Ultimately, we expect to identify the culprit gene(s) and synaptic targets that cause TC abnormalities and abnormal cortical activity in these mouse models of SCZ. This information will provide a framework for the future development of specific therapeutic interventions to alleviate positive symptoms in patients with this catastrophic disease.
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1 |
2021 |
Reiter, Lawrence T [⬀] Zakharenko, Stanislav S |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
The Role of Ube3a in Gliopathic Seizures @ University of Tennessee Health Sci Ctr
As many as ~3 million individuals suffer from epileptic seizures in the US alone. Although some forms of epilepsy respond quite well to anti-seizure medications, a substantial portion of individuals suffering from epilepsy do not respond well to medication or dietary treatments. One way to investigate the underlying molecular and cellular pathology of seizure disorder is by studying a genetically defined syndrome where epilepsy is a prominent feature. Approximately 25-50% of individuals with Duplication 15q syndrome (Dup15q) suffer from difficult to control seizures. The prevailing hypotheses to explain seizures in Dup15q are maternal specific neuronal elevation of the ubiquitin E3 ligase UBE3A and/or duplication of a cluster of GABA receptor genes located adjacent to UBE3A at 15q11.2-q13.1. Studies using the fruit fly, Drosophila melanogaster, from our laboratory indicates that these seizures may be caused by elevated levels of UBE3A in glia, not neurons as previously proposed. The premise for this proposal is that seizures are caused by elevated levels of the UBE3A protein in glia, not neurons, providing not only a pathway to molecular mechanism for Dup15q related epilepsy, but also a paradigm shift, directly implicating glial cells in the etiology of seizures. The experiments outlined in this proposal are designed to investigate the developmental timing and molecular changes in glial cells over-expressing Dube3a and to reveal how these changes are recapitulated in a new mouse model we developed that expresses Ube3a in glial cells. Everything we learn from these studies in flies will be directly disseminated to a team of seizure experts who work at the Duplication 15q centers of excellence throughout the country. The identification of new therapeutic targets for Dup15q epilepsy may also provide new treatment options to individuals who are pharmacoresistant to current anti-epileptics.
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0.912 |