Lynn Raymond

Professor

Relevant Degree Programs

 

Graduate Student Supervision

Doctoral Student Supervision (Jan 2008 - May 2019)
Global neuroprotection of huntingtin in culture and alterations of cortico-striatal connections in Huntington’s disease culture and mouse models (2017)

Huntington’s disease (HD) is a genetic neurodegenerative disorder caused by expansion of a CAG repeat in exon 1 of the HTT gene, encoding an elongated poly-glutamine repeat in the N-terminal region of the protein huntingtin (mutant huntingtin; mHtt). The average age of onset is 38, and the disease is characterized by psychiatric disorders and cognitive deficits that, in general, gradually develop over 10 years before the overt onset of the disease phenotype – difficulties in movement control. In the past two decades, many studies have focused on cell death that is obvious in mid to late stage of the disease when the overt disease symptoms become irreversible, despite the fact that altered neuronal/synaptic functions may underlie the mood/cognitive disorders that precede a motor diagnosis.In order to uncover the potentially preventable and/or reversible changes in cortico-striatal (C-S) connections in pre- and early stages of HD, we first studied the C-S coculture platform that represents its in vivo counterparts in order to investigate the role of wild-type huntingtin (wtHtt) protein in cell-death and C-S synaptic malfunctions in HD. I found that coculture with low cortical-to-striatal neuronal plating ratio (1:3 plating ratio) is a closer replica of its in vivo origin with slight differences in membrane properties, but with a significant increase in extrasynaptic NMDA receptor portion and a decrease in cell-survival signaling compared with the control (1:1). On the other hand, we found that wtHtt provides neuroprotective effects to striatal, cortical and hippocampal neurons, in a phospho-CREB-independent way in the case of the latter two neuronal types. Finally, using the C-S coculture and acute brain slice to study C-S synapse development and functions, I found that mHtt impairs the connection not only via suppressing striatal dendritic tree development but also by altering excitatory presynaptic vesicle release and recovery of the glutamate pool.In summary, this work is a further proof of HD as synaptopathy, and is a foundation for future research of drug discovery for HD targeting synaptic malfunctions at the pre-symptomatic stage.

View record

Signaling pathways involved in enhanced NMDA receptor-dependent excitotoxicity in a mouse model of Huntington disease (2011)

Huntington disease (HD) is an inherited neurodegenerative disease lacking effective treatment, characterized by involuntary movements, psychiatric disorders, and cognitive symptoms. Pathology shows prominent degeneration of γ-aminobutyric acid (GABA)-ergic medium-sized spiny neurons (MSNs) of the striatum and certain cortical layers (Vonsattel and DiFiglia, 1998). HD is caused by a dominant mutation in the HD gene that leads to >35 glutamine repeats (polyQ) near the N-terminus of the protein huntingtin (htt) (The Huntington’s Disease Collaborative Research Group, 1993). Increasing evidence suggests that the N-methyl-D-aspartate (NMDA)-type glutamate receptor (NMDAR) plays a role in mediating death of MSNs observed in HD (Fan and Raymond, 2007). Previous results from our laboratory demonstrate that NMDAR-mediated current and toxicity are increased in MSNs from the Yeast Artificial Chromosome (YAC) transgenic mouse model expressing polyglutamine-expanded full-length human htt (Shehadeh et al., 2006; Zeron et al., 2002). However, the mechanism underlying altered function and enhanced toxicity of NMDAR in HD remains unknown. Previous studies have shown that membrane-associated guanylate kinases (MAGUKs), such as postsynaptic density protein 95 (PSD-95) modulate NMDAR surface expression and excitotoxicity in rat hippocampal and cortical neurons (Aarts et al., 2002; Roche et al., 2001), and that htt interacts with PSD-95 in a polyglutamine dependent manner (Sun et al., 2001). Here, I tested the hypothesis that an altered association and/or regulation between PSD-95 and NMDARs in mutant htt-expressing cells contributes to increased susceptibility to excitotoxicity and investigated mechanism by which this occurs. Specifically, I investigated the association of PSD-95 with htt and the NMDAR GluN2 subunits; signaling downstream of activation of the NMDAR/PSD-95 complex; and NMDA-induced cell death. My results suggest that at the presymptomatic stage of HD, the enhanced interaction of PSD-95 with GluN2B, and its signaling through p38 mitogen-activated protein kinase (MAPK) but not neuronal nitric oxide synthase (nNOS) activation, contributes to mutant htt-mediated sensitivity to NMDAR-dependent excitotoxicity in YAC128 striatal neurons. This work contributes to the understanding of both NMDAR-dependent neuronal death mechanisms in striatal neurons and early synaptic changes in HD pathogenesis, as well as providing potential drug candidates for future HD treatment.

View record

Functional regulation of N-methyl-D-aspartate receptor subtypes and their involvement in hippocampal plasticity (2010)

Regulation of NMDAR activity by desensitization is important in physiological and pathological states. We previously reported that desensitization decreases during hippocampal neuronal development, correlating with NMDAR composition, synaptic localization and association with PSD-95. To determine if PSD-95-induced changes in NMDAR desensitization occur because of direct binding to NR2 subunits or due to recruitment of regulatory proteins, we tested the effects of various PSD-95 constructs on NMDAR currents in HEK293 cells and neurons. In HEK cells, wt PSD-95 significantly reduced wt NMDAR desensitization without altering currents of NMDARs containing NR2A-S1462A, a mutation that abolishes PSD-95 binding. Moreover, PDZ1-2 domain was sufficient for this effect in neurons with low endogenous PSD-95 levels. Moreover, other PSD-95 family members with highly homologous PDZ1-2 domains significantly reduced NMDAR desensitization. In mature neurons, disruption of PSD-95/NMDAR interaction through PKC activation, or through interference peptides, increased desensitization to levels found in immature neurons. We conclude that direct binding of PSD-95 increases stability of NMDAR responses to agonist exposure.Desensitization is a property that shapes synaptic responses, and modulates the calcium signal mediated by the two predominant NMDARs subtypes in hippocampus, with possible consequences for their functioning. Further, we examined the involvement of NR2 subtypes in synaptic plasticity in hippocampal dentate gyrus of juvenile mice. Exercise was used as a means to alter expression of the NR2 subunits in this region. We compared two groups of animals: Controls, which were housed in conditions of minimal enrichment, andiiiRunners, which had access to an exercise wheel. NMDAR-dependent LTP expression was significantly greater in Runners than in Controls; in the presence of NR2B subunit antagonists, it was significantly reduced in both groups. NR2A subunit antagonist blocked LTP in slices from Runners and produced a slight depression in Control animals. LTD could not be prevented by either of the NR2B specific antagonists. Strikingly, eliminating NR2A subunit-containing receptor activity prevented LTD in Runners, but not in Control animals. Overall, these results indicate that interplay between subtype, subcellular localization and size of NMDAR subpopulations accounts for their diverse role in synaptic plasticity induction, and that exercise increases the contribution of NR2A to plasticity.

View record

Master's Student Supervision (2010 - 2018)
Palmitoylation on NMDA receptor trafficking and function in corticostriatal co-culture from HD mouse model (2016)

Huntington’s Disease (HD) is a neurodegenerative disorder in which the medium-sized spiny neurons (MSNs) of the striatum are earliest and most severely affected. Selective striatal degeneration in HD has been caused, in part, by altered N-methyl-D-aspartate receptor (NMDAR) activity. Our lab has found in the YAC128 transgenic mouse model of HD that GluN2B-containing NMDARs (2B-NMDARs) at extrasynaptic (Ex) sites are increased in striatum at an early stage; however, the mechanism underlying altered NMDAR trafficking in HD remains unknown. Palmitoylation at the C-terminus of 2B-NMDAR regulates its surface expression and synaptic targeting. Notably, a palmitoyl transferase (PAT) enzyme – ZDHHC17 (HIP14) – interacts with huntingtin, the protein mutated in HD. Pilot data in the lab suggests reduced 2B-NMDAR palmitoylation may contribute to increased Ex-NMDAR in striatal MSN from YAC128 HD mice. However, a potential role for HIP14 in regulating 2B-NMDAR trafficking has not been explored. On the other hand, suppression of acyl-protein thioesterases 1/2 (APT1/2), the depalmitoylation enzymes, by a small molecule inhibitor PalmB leads to increased protein palmitoylation level, which may affect NMDAR distribution. Here, we examined the effect of changing palmitoylation on NMDAR trafficking and function in YAC128 HD mice. Knockdown of endogenous HIP14 did not change 2B-NMDAR surface expression and total NMDAR current. The treatment of DMSO (vehicle) and PalmB reduced synaptic NMDAR current and increased Ex-NMDAR current, and the main difference was found in FVB/N (control) mouse. In contrast, after 4 hours of PalmB treatment, miniature excitatory postsynaptic current (mEPSC) frequency significantly increased at YAC128 but not FVB/N corticostriatal synapse. Investigation of palmitoylation on NMDAR activity is useful for clinical application.

View record

Coupling extrasynaptic NMDA receptors to aberrant intracellular signaling in Huntington disease (2013)

N-methyl-D-aspartate glutamate receptors (NMDARs) play dichotomous roles on neuronal survival, depending on their surface localization: while synaptic NMDARs promote pro-survival pathways, those expressed at extrasynaptic sites (Ex-NMDARs) trigger pro-death cascades. In the YAC128 transgenic mouse model of Huntington disease (HD), elevated Ex-NMDAR expression contributes to the onset of cognitive dysfunction and striatal death. A shift in the balance of synaptic-extrasynaptic NMDAR signaling and localization is paralleled by dysregulation of intracellular calcium signaling pathways that couple to pro-death cascades. However, whether aberrant calcium signaling is a consequence of elevated Ex-NMDAR expression in HD is unknown. Here, we examined calcium-dependent pathways downstream of Ex-NMDARs in HD. Chronic (2-month) treatment of YAC128 and WT mice with memantine (1 and 10mg/kg/d), which at a low dose selectively blocks Ex-NDMARs, reduced striatal Ex-NMDAR expression in YAC128 mice without altering synaptic NMDAR levels. In contrast, calpain activity was not affected by memantine treatment, and was elevated in untreated YAC128 mice at 1.5 months but not 4 months of age. In YAC128 mice, memantine at 1mg/kg/d rescued CREB shut-off, while both doses suppressed p38 MAPK activation to WT levels. In contrast, extrasynaptic PSD-95 expression was not affected by memantine in YAC128 mice but was increased by memantine at 10mg/kg/d in WT littermates. Hence, Ex-NMDAR activity drives increased extrasynaptic receptor expression as well as dysregulated p38 MAPK and CREB signaling in HD. Elucidation of the pathways centered around Ex-NMDARs in HD could help provide novel therapeutic targets for this disease.

View record

Opposing roles of synaptic and extrasynaptic NMDA receptor signaling in co-cultured striatal and cortical neurons (2011)

The N-methyl-d-aspartate receptor- type glutamate receptor (NMDAR) plays a unique and vital role in subcellular signaling. Ca²⁺ influx initiates signaling cascades important for synaptic plasticity and survival. However, overactivation of the receptor leads to toxicity and cell death. This dichotomy is partially explained by the subcellular location of the receptor. NMDARs located at the synapse have been shown to signal for cell survival, while extrasynaptic receptors signal for cell death. Thus far, the interplay between synaptic and extrasynaptic NMDARs has been studied exclusively in cortical (CTX) and hippocampal neurons; it is unknown whether medium spiny neurons of the striatum (MSNs), which bear the brunt of neurodegeneration in Huntington disease, follow the same pattern. There is evidence to suggest that signaling pathways may be different in CTX compared with MSNs. Here we study, for the first time, synaptic versus extrasynaptic signaling in striatal MSNs, focusing on activation of cAMP response element binding protein (CREB). Synaptic NMDARs activate CREB in striatal MSNs, although this pathway is slightly less efficacious compared with CTX. Similarly to CTX, extrasynaptic NMDARs shut off CREB in MSNs. MSNs are less susceptible to NMDA-mediated toxicity compared with CTX. Blocking extrasynaptic receptors with memantine (30 µM) and GluN2B-containing receptors with ifenprodil (3 µM) prevents CREB shutoff and rescues neurons from NMDA-mediated toxicity. This work may provide cell- and NMDAR subtype-specific targets for treatment of diseases with putative NMDAR involvement, including neurodegenerative diseases and ischemia.

View record

Current Students & Alumni

This is a small sample of students and/or alumni that have been supervised by this researcher. It is not meant as a comprehensive list.
 
 

If this is your researcher profile you can log in to the Faculty & Staff portal to update your details and provide recruitment preferences.