Synaptic and extrasynaptic localization of brain-derived neurotrophic factor and the tyrosine kinase B receptor in cultured hippocampal neurons.
Brain-derived neurotrophic factor (BDNF) regulates synapses, but the distribution of BDNF and its receptor TrkB relative to the location of glutamatergic and gamma-aminobutyric acidergic (GABAergic) synapses is presently unknown. Immunocytochemistry was performed in primary hippocampal neuron cultures to determine whether BDNF and TrkB are preferentially localized to excitatory or inhibitory markers at 7, 14, and 21 days in vitro (DIV). Glutamatergic sites were localized with vesicular glutamate transporter type 1 (VGLUT1) as presynaptic marker and the NR1 subunit of the NMDA receptor and the GluR1 subunit of the AMPA receptor as receptor markers. GABAergic sites were labeled with the 65-kDa isoform of glutamic acid decarboxylase (GAD-65) as presynaptic marker and the gamma2 subunit of the GABAA receptor as receptor marker. During development, <30% of BDNF punctae and TrkB clusters were localized to glutamatergic and GABAergic markers. Because their rates of colocalization did not change from 7 to 21 DIV, this study details the distribution of BDNF and TrkB at 14 DIV. BDNF was preferentially colocalized with glutamatergic markers VGLUT1 and NR1 ( approximately 30% each). TrkB was also relatively highly colocalized with VGLUT1 and NR1 ( approximately 20% each) but was additionally highly colocalized with GABAergic markers GAD-65 ( approximately 20%) and gamma2 ( approximately 30%). NR1 clusters colocalized with BDNF puncta and TrkB clusters were mostly extrasynaptic, as were gamma2 clusters colocalized with TrkB clusters. These results show that, whereas most BDNF and TrkB protein is extrasynaptic, BDNF is preferentially associated with excitatory markers and that TrkB is associated equally with excitatory and inhibitory markers.
Swanwick
Catherine Croft
CC
Neuroscience Graduate Program, University of Virginia, Charlottesville, Virginia 22908, USA.
Harrison
Madaline B
MB
Kapur
Jaideep
J
D000818:Animals; D019208:Brain-Derived Neurotrophic Factor / Q000378:metabolism* / Q000502:physiology; D002478:Cells, Cultured; D006624:Hippocampus / Q000737:chemistry / Q000254:growth & development / Q000378:metabolism*; D009474:Neurons / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D020813:Receptor, trkB / Q000378:metabolism*; D013569:Synapses / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology
D019208:Brain-Derived Neurotrophic Factor; D020813:Receptor, trkB