1,721,090 research outputs found
Native glycine receptor subtypes and their physiological roles
No full textThe glycine receptor chloride channel (GlyR), a member of the pentameric Cys-loop ion channel receptor family, mediates inhibitory neurotransmission in the spinal cord, brainstem and retina. They are also found presynaptically, where they modulate neurotransmitter release. Functional GlyRs are formed from a total of five subunits (alpha 1 -alpha 4, beta). Although alpha subunits efficiently form homomeric GlyRs in recombinant expression systems, homomeric alpha 1, alpha 3 and alpha 4 GlyRs are weakly expressed in adult neurons. in contrast, alpha 2 homomeric GlyRs are abundantly expressed in embryonic neurons, although their numbers decline sharply by adulthood. Numerous lines of biochemical, biophysical, pharmacological and genetic evidence suggest the majority of glycinergic neurotransmission in adults is mediated by heteromeric alpha 1 beta GlyRs. Immunocytochemical co-localisation experiments suggest the presence of alpha 2 beta, alpha 3 beta and alpha 4 beta GlyRs at synapses in the adult mouse retina. Immunocytochemical and electrophysiological evidence also implicates alpha 3 beta GlyRs as important mediators of glycinergic inhibitory neurotransmission in nociceptive sensory neuronal circuits in peripheral laminae of the spinal cord dorsal horn. It is yet to be determined why multiple Glyl? synaptic subtypes are differentially distributed in these and possibly other locations. The development of pharmacological agents that can discriminate strongly between different beta subunit-containing GlyR isoforms will help to address this issue, and thereby provide important insights into a variety of central nervous system functions including retinal signal processing and spinal pain mechanisms. Finally, agents that selectively potentiate different GlyR isoforms may be useful as therapeutic lead compounds for peripheral inflammatory pain and movement disorders such as spasticity. (C) 2008 Elsevier Ltd. All rights reserved
Determining the Role of the Complement Cascade in Hippocampal Synaptic Physiology
Full text linkA key feature of brain development is the refinement of exuberant synapses, known as synapse pruning. This phenomenon is executed by microglia, the resident immune cells of the brain. Evidence that microglia prune retinal inputs via activation of the complement cascade in the dorsolateral geniculate nucleus has led to the principle that complement-dependent synapse pruning occurs ubiquitously in the brain. However, this principle has not been empirically validated. Here, I investigated this principle in the hippocampus, a critical region for learning and memory which is known to undergo developmental pruning by microglia. In the healthy brain, complement cascade proteins are enriched in VGLUT2+ regions of the hippocampus. Therefore, I hypothesized that in the developing hippocampus, microglia-mediated pruning of VGLUT2+ synapses is complement-dependent. To test this hypothesis, I used mice with a genetic deletion of the essential complement protein C3. Microglia pruning of VGLUT2+ synapses was not different between C3 -/- mice compared to WT during development in the hippocampus. Further, functional synapse properties and plasticity were not significantly altered in C3 -/- compared to WT mice. Thus, complement-dependent pruning by microglia is a not general mechanism in the brain under physiological conditions. I then tested whether a complement-dependent mechanism could be induced in the hippocampus by peripheral inflammation. I found that treating mice systemically with lipopolysaccharide (LPS) led to a neuroinflammatory microglia phenotype and VGLUT2+ synapse loss. Surprisingly, the neuroinflammatory microglia phenotype was exacerbated in LPS-treated C3 -/- mice compared to LPS-treated WT mice. As such, in an inflammatory context, complement C3 acts as a negative regulator of microglia reactivity in the hippocampus. Finally, I observed that under physiological conditions, an immature-like phagocytic state persists in adulthood specifically in the CA1 SLM, but not in adjacent CA1 SR and DGML hippocampal subregions. Together, my Thesis work has i) provided a counterexample to the principle of ubiquitous complement-dependent synapse pruning by microglia, ii) uncovered an unexpected role for C3 in neuroinflammation, and iii) revealed that phagocytic activity in the hippocampus is governed in a subregion-dependent manner. These findings refine our understanding of microglia function in the hippocampus.Ph.D.2024-11-13 00:00:0
A Novel Role of STAT3 in Synaptic Plasticity in Juvenile Mice
Full text linkThe dynamic and complex process by which neurons modulate their connections is termed synaptic plasticity. Synaptic connections between neurons can be strengthened or weakened in an activity-dependent manner, through long-term potentiation (LTP) or long-term depression (LTD), respectively. In recent years, the Janus Kinase (JAK) and Signal Transducer and Activator of Transcription (STAT) signaling pathway, traditionally involved in immune signaling, was implicated in synaptic plasticity. More specifically, STAT3 was required for N-methyl-D-aspartate receptor (NMDAR)-LTD in distinct hippocampal CA1 synapses in juvenile and adult rats. Studies in rats utilized pharmacological and knockdown approaches to evaluate STAT3 in LTD. This thesis aimed to further elucidate the role of STAT3 in LTD and in additional forms of plasticity through conditional genetic targeting and pharmacological inhibition in mice across development.Two brain-specific conditional STAT3 deletion mouse lines were generated through CaMKII- and Emx1-mediated Cre expression, allowing for investigations into hippocampal NMDAR-dependent synaptic plasticity in juvenile and adult mice. Conditional knockout (KO) and pharmacological inhibition of STAT3 did not impair basal synaptic transmission or synaptic facilitation in either age cohort studied. Furthermore, adult CaMKII-STAT3 KO mice, juvenile Emx1-STAT3 KO mice, and juvenile wildtype mice treated with pharmacological inhibitors of STAT3 displayed intact NMDAR-LTD. Two theta burst stimulation (TBS) protocols, either in a compressed (cTBS) or spaced (sTBS) pattern, were used to study STAT3 in LTP. These protocols trigger distinct forms of LTP reliant on different postsynaptic expression mechanisms. Juvenile Emx1-STAT3 KO mice displayed a significant reduction in LTP following a cTBS, but not an sTBS induction protocol. Adult CaMKII-STAT3 and adult Emx1-STAT3 KO mice displayed normal LTP following either protocol. This LTP deficit in juvenile KOs following cTBS was linked to protein kinase A (PKA) activity. Inhibition of PKA significantly increased LTP in Emx1-STAT3 KOs despite cTBS LTP being PKA-independent in Emx1-STAT3 controls. Taken together, I identified a novel developmental requirement for STAT3 in a specific form of LTP following cTBS. This work highlights the importance of comprehensive developmental studies and emphasizes the complexity of synaptic plasticity in the CA1 region in mice.
Key words: STAT3, synaptic plasticity, CA1, long-term potentiation, long-term depression, protein kinase A, juvenile micePh.D.2025-11-13 00:00:0
Role of Intracellular Calcium Stores and Calcium-Permeable AMPARs in Functional and Structural Plasticity in Mouse Hippocampal Slices
Full text linkMemories are internal representations of life that can last in the order of minutes to years, and are largely experience-dependent. For example, the memory of an inconsequential event typically fades, unless they are closely related to a novel event, succinctly explained by the synaptic tag and capture (STC) hypothesis. The formation of short (STM)- or long - term memories (LTM) involve several biochemical changes that either increase or decrease synaptic efficacy. These changes are commonly referred to as Long-term potentiation (LTP) or depression (LTD). LTP can be differentiated into two forms based on their dependence (LTP2) or independence (LTP1) on protein synthesis (PS) and are hypothesized to be analogous to LTM and STM respectively. In both forms of LTP, calcium (Ca2+) signaling plays a vital role. However, Ca2+ has a number of different sources, the relative importance of which is poorly understood. Here I have investigated
the role of calcium-permeable (CP)- AMPARs and Ca2+ release from intracellular stores. Using theta burst stimulation (TBS) of varied strength and patterning, I was able to selectively induce LTP1 (wTBS or cTBS) or a combination of LTP1 and 2 (sTBS). Combining these induction protocols with pharmacological agents, I have confirmed the role of CP-AMPARs and demonstrated that intracellular Ca2+ stores are required specifically for LTP2. To investigate STC, I delivered LTP 2 and LTP1 to independent inputs, which resulted in an enhancement of LTP1 due to STC. I confirmed the role of CP-AMPARs and demonstrated the importance of Ca2+ release from intracellular stores during the induction of LTP2 in STC.
Furthermore, to address the question of the association between functional and structural plasticity under physiological conditions, I developed a method to monitor both simultaneously using field potential recordings and two-photon imaging, respectively. I found that functional plasticity induced by sTBS is associated with structural plasticity and that the structural plasticity is entirely dependent on the activation of CP-AMPARs. In conclusion, my findings provide novel insights to the synaptic basis of memory formation and highlight the importance of CP-AMPARs and calcium release from intracellular stores in this process.Ph.D
Alterations in Synaptic Plasticity in a Mouse Model of Fragile X Syndrome
Full text linkFragile X Syndrome (FXS), a form of inherited intellectual disability that is caused by a mutation in the Fragile X Messenger Ribonucleoprotein 1 (FMR1) gene, is commonly studied using the Fmr1 knockout (KO) mouse model. Studies in Fmr1 KO mice have demonstrated altered synaptic plasticity, — dysfunctional up- and down-regulation of synaptic strength — synaptic potentiation and depression, respectively. Early interventions in FXS focussed on metabotropic glutamate receptor (mGluR) long-term depression, which is enhanced in Fmr1 KO mice. However, inhibitors of mGluRs, which normalise FXS phenotypes in pre-clinical models, were unsuccessful in clinical trials. Deficits in N-methyl-D-aspartate receptor (NMDAR) long-term potentiation (LTP) have also been identified in the Fmr1 KO mouse, however there are discrepancies in the literature when it comes to the severity of LTP phenotypes. Meanwhile, NMDAR-dependent short-term potentiation (STP) deficiencies have not been researched in the Fmr1 KO mouse.
Chapter 1 describes the background of this thesis, while Chapter 2 details the methods used during the investigations in Chapters 3 – 5. Chapter 3 investigated whether the breeding strategy (termed cage effect) affects LTP, STP, contextual fear conditioning (CFC) and glutamate receptor expression in ten groups of male and female Fmr1 mice, bred and maintained in different ways. Only male non-littermate KO mice had deficits in STP, LTP and CFC. Expression of glutamate receptors was unaffected by genotype, but was strongly influenced by the cage effect in male littermate WT and KO mice. Chapter 4 demonstrated that the LTP deficiency, observed in the non-littermate KO mice, can be reversed by potentiation of NMDARs; by using a positive allosteric modulator, UBP714, or a partial NMDAR agonist, D-Cycloserine. Finally, Chapter 5 investigated the extent to which induction of STP and LTP differed between non-littermate WT and KO mice by using induction protocols of increasing strength and found that KOs have consistently smaller STP, but a higher threshold for LTP saturation than the WTs. Plasticity in KO mice showed increased sensitivity to UBP791, an NMDAR antagonist. Overall, this thesis highlights that cage effects, induction protocols and sex differences can influence phenotypes in Fmr1 colonies and may explain the phenotype discrepancies in the field.Ph.D
Investigating the Role of LIMK1 Signaling in PKA-dependent LTP in the Hippocampus
No full textLong-term potentiation (LTP) in the hippocampus is the most extensive form of long-lasting synaptic plasticity and is widely regarded as the cellular basis of memory formation. The canonical NMDA-receptor (NMDAR)-dependent form of LTP can be differentiated into protein kinase A (PKA)-dependent and -independent forms by the spacing between theta burst stimuli (TBS) induction. Key features of PKA-dependent LTP includes insertion of Ca2+ permeable AMPA receptors (CP-AMPARs) and initiation of de novo protein synthesis. However, the molecular mechanisms that elicit these processes remain unknown. Previous studies suggest PKA can regulate LIM-domain kinase (LIMK) 1, but this regulation has not been demonstrated in synaptic plasticity. Using a combination of electrophysiology, genetic mouse models, and pharmacology, I show that CP-AMPAR-dependent LTP requires PKA regardless of TBS spacing. Furthermore, I demonstrate that LIMK1 is required for PKA-dependent LTP. These findings provide insight into the molecular mechanisms underlying LTP and long-term memory formation.M.Sc.2020-03-21 00:00:0
NMDA Receptor Hypofunction in the Mouse Anterior Cingulate Cortex Associated with a De Novo GRIN1 Mutation at the M2 Reentrant Loop
Full text linkThe dysfunction of N-methyl-D-aspartate receptors (NMDARs) is consistently observed in a wide range of neurodevelopmental disorders and neurodegenerative diseases. Mutations of the GluN1 subunit of NMDARs have been shown to impair receptor trafficking and function which is thought to contribute to deficits in cognitive function observed in GRIN1-related neurodevelopmental disorders. Other clinical characteristics of GRIN1 variants such as altered pain sensitivity highlight a possible role of the anterior cingulate cortex (ACC), although the effects of GRIN1 variants at excitatory ACC synapses have not yet been studied. Whole-cell electrophysiology was used to examine NMDAR-mediated synaptic transmission and the intrinsic membrane properties of ACC layer II/III pyramidal neurons. The presence of the GRIN1_G620R missense mutation was associated with significant reductions in evoked NMDAR-mediated synaptic transmission, as well as changes to membrane input resistance and capacitance suggesting altered neuronal morphology. Our findings indicate that the ACC, a major region implicated in the processing of pain and emotion, is severely affected by the GRIN1_G620R missense variant in regard to the impairment of excitatory synaptic transmission.M.Sc
The Effects of Alzheimer's Disease on Synaptic Plasticity and the cAMP/PKA Pathway
Full text linkAlzheimer’s Disease (AD) is a physiologically devastating form of dementia, yet an understanding of AD pathology remains elusive. One promising avenue of research is to examine synaptic plasticity within the brain centres of learning and memory in animal models of AD. Long-term potentiation (LTP) is a model of learning and memory at the synaptic level, and deficits in LTP have been proposed in AD. The aim of this work is to examine whether synaptic defects in AD involve a neuronal signalling cascade: the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) pathway.
This research uses novel modes of LTP induction which manipulate the course of LTP induction. Examinations reveal alterations in basal synaptic transmission in AD mouse models. The use of a cAMP/PKA enhancing drug revealed a potential dysregulation in the cAMP/PKA cascade. Lastly, coastline analysis showed potentiation induced hyperexcitability at synapses of an AD mouse model.M.Sc
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