The Role of PSD-95 and Cypin in Morphological Changes in Dendrites Following Sublethal NMDA Exposure

Abstract

Focal swelling or varicosity formation in dendrites and loss of dendritic spines are the earliest indications of glutamate-induced excitotoxicity. Although it is known that microtubule dynamics play a role in varicosity formation, very little is known about the proteins that directly impact microtubules during focal swelling and dendritic spine loss. Our laboratory has recently reported that the postsynaptic protein PSD-95 and its cytosolic interactor (cypin) regulate the patterning of dendrites in hippocampal neurons. Cypin promotes microtubule assembly, and PSD-95 disrupts microtubule organization. Thus, we hypothesized that cypin and PSD-95 may play a role in altering dendrite morphology and spine number in response to sublethal NMDA-induced excitotoxicity. Using an in vitro model of glutamateinduced toxicity in rat hippocampal cultures, we found that cypin overexpression or PSD-95 knockdown increases the percentage of neurons with varicosities and the number of varicosities along dendrites, decreases the size of varicosities after sublethal NMDA exposure, and protects neurons from NMDA-induced death. In contrast, cypin knockdown or PSD-95 overexpression results in opposite effects. We further show that cypin regulates the density of spines/filopodia: cypin overexpression decreases the number of protrusions per micrometer of dendrite while cypin knockdown results in an opposite effect. Cypin overexpression and PSD-95 knockdown attenuate NMDA-promoted decreases in protrusion density. Thus, we have identified a novel pathway by which the microtubule cytoskeleton is regulated during sublethal changes to dendrites.

Autism-specific maternal autoantibodies recognize critical proteins in developing brain

“…The number of protrusions per dendrite decreased with an overexpression of cypin, whereas a cypin knockdown resulted in an increase in spines/filopodia on the dendrite. 35 In another study, exposure to brain-derived neurotrophic factor, an important mediator of dendrite arborization and a factor previously implicated in ASD, increases cypin in cultured rat hippocampal neurons. Thus, one can speculate that interfering with cypin through autoantibody exposure during neurodevelopment could affect dendrite formation, potentially increasing the number of dendritic spines in certain regions of the brain.…”

Glutamate affects dendritic morphology of neurons grown on compliant substrates

“…Although debated, the concentration of glutamate in the brain is believed to be in the micromolar range Synaptic cleft concentrations are thought to be on the order of 150–200 µM . Our studies included treatment of neurons with concentrations of 10 µm glutamate, within the ambient concentration in the brain and at a much lower level that what we have determined to be neurotoxic in our cultures . Thus, even if glutamate levels are not released at toxic amounts after injury, ambient glutamate itself may modulate effects of the injury.…”

“…34 Our studies included treatment of neurons with concentrations of 10 mm glutamate, within the ambient concentration in the brain and at a much lower level that what we have determined to be neurotoxic in our cultures. 35 Thus, even if glutamate levels are not released at toxic amounts after injury, ambient glutamate itself may modulate effects of the injury.…”

The Nuclear Calcium Signaling Target, Activating Transcription Factor 3 (ATF3), Protects against Dendrotoxicity and Facilitates the Recovery of Synaptic Transmission after an Excitotoxic Insult

“…Although calpain can cleave microtubules, this calcium-activated protease is unlikely to be involved in initial dendritic beading (30,53). However, both dendritic beading and microtubule fragmentation are prevented by pretreatment with the microtubule-stabilizing compound taxol (34,(53)(54)(55)(56). These findings suggest that both the initial disruption of microtubule morphology and dendritic beading may result from excessive water influx.…”