UCSD.EDU. Introduction Importance of Synapses -brain development -crucial for brain function.

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Presentation transcript:

UCSD.EDU

Introduction Importance of Synapses -brain development -crucial for brain function

THE GLUTAMATERGIC SYNAPSE Presynaptic Postsynaptic Andrea Draper

Why Neuroligin1 vs. Other Proteins? WADSWORTH CENTER Synaptogenic Interacts with key post synaptic protein (PSD95) Knock-out mice reveal necessary for survival Tentative link to neuronal developmental disorders

Hypothesis NLG1 may recruit the scaffolding proteins PSD95, SAP102, and S-SCAM to the post synaptic density via different intracellular domains.

Experimental Strategies If I get rid of the PDZ domain of NLG1, will that have an effect on the localization of post synaptic proteins? Answer: Over express NLG1 and NLG1-d55 DNA in neurons Stain neurons Quantify and compare normal neurons to DNA over expressed neurons Question?

Goal Observe the effect of manipulating the NLG1 structure

Neurons are extracted from hippocampus and cultured 1) Neurons are either stained or transfected and then stained 2) Experimental Procedures

IMMUNOCYTOCHEMISTRY SAP102PSD95 CO-LOCALIZATION

Transfected Neuron Control Co-localization of pre and post synaptic proteins

Experiment: Manipulating NLG1 HA PDZ HA-NLG1 HA-NLG1-d55 vs. HA PDZ HA PDZ

Antibody Staining of Transfected Neurons HA-NLG1 Co-localization of pre and post synaptic proteins HA-NLG1PSD95 V-GLUT PSD95

Antibody Staining of Transfected Neurons HA-NLG1-d55 Co-localization of pre and post synaptic proteins

Results Control NLG1-d55 TransfectionNLG1 Transfection

Results My lab has found previously that by manipulating the PDZ domain of NLG1, it is found that this does have an effect on the co-localization at a synapse.

Hypothetical Results HA AChE PDZ Increased # synapse HA AChE Decreased # synapse HA-NLG1 HA-NLG1-d55 Untransfected Neuron vs. NLG1 and d55

Observe effect of NLG1 manipulation on the localization of post synaptic proteins Goal

Future Directions -Test NLG1 difference in younger vs. older neurons -Effects of NLG1 on localization of other post synaptic proteins -NLG1 linked to neuronal developmental disorder?

Acknowledgements -Jennifer Hoy -Phil Washbourne and the lab -SPUR program -SPUR students -University of Oregon