Probing the ability of presynaptic tyrosine kinase receptors to regulate striatal dopamine dynamics.

Neurotrophic factors are endogenous soluble proteins that regulate the cell cycle, growth, differentiation, and survival of neurons.1 Members of the neurotrophic family include nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), glial-derived neurotrophic factor (GDNF), neurotrophin-3 (NT-3), and neurotrophin-4 (NT-4), all of which mediate their functions through tyrosine kinase (Trk) receptors. The Trk receptors are glycoproteins that have a molecular weight in the range of 140–145 kDa. Each neurotrophin appears to bind to a unique isoform of the Trk receptors. For example, NGF has a greater specificity to bind to the TrkA receptor, NT-3 interacts with TrkC, and both BDNF and NT-4 bind to TrkB.2,3 When BDNF is expressed, it is transported to nerve terminals through the axon and undergoes exocytotic release from presynaptic vesicles.4−6 Extracellular BDNF binds to TrkB receptors and causes receptor dimerization, which leads to phosphorylation of tyrosine residues within the cytoplasm, and activates kinases.7 Phosphorylated tyrosine residues recruit specific proteins, such as GBR2, SHC, and SOS, which in turn activate Raf, a serine/threonine protein kinase (Akt).8 Trk activation induces numerous signaling cascades through three known pathways: the Ras/mitogen activated protein kinase (MAPK), phosphatidylinositol-3-kinase (PI3K), and phospholipase C, γ (PLCγ). Activating these signaling cascades regulate the transcription factor, cAMP response element binding (CREB), and leads to attenuation or potentiation in gene expression. Continuous activation or inhibition of these signaling events regulate synaptic plasticity, synaptic transmission, neurotransmitter release, neurogenesis, and cell survival.3,6,9−11 There is considerable evidence of reciprocal interactions between BDNF and the dopamine (DA) system, although the exact mechanism of how BDNF regulates DA dynamics is unknown.11−13 BDNF regulates striatal function directly by activating TrkB receptors.14 When BDNF is infused directly into the brain it influences the survival and function of DA neurons, affects the turnover ratio between DA and its metabolite DOPAC, and potentiates the activity-dependent release of DA.11−13 Evidence suggests that BDNF expression can augment DA transmission in the reward pathway of the ventral tegmental-nucleus accumbens (NAc) circuit.15 Mouse models with reduced BDNF expression exhibit a variety of alterations in the DA system, which indicate that BDNF has some influence on this system. Studies have shown that mice lacking one copy of the BDNF gene (BDNF+/–) have higher tissue DA concentrations in the striatum, and decreased DA release in superfused striatal tissue fragments.16,17 BDNF knockout (BDNF–/–) and BDNF+/– mice have reduced expression and density of DA D3 receptors in the caudate-putamen (CPu), NAc core and shell, and in the Islands of Calleja.18−20 Our laboratory has shown that BDNF+/– mice have an ∼2.5-fold increase in extracellular DA levels in the CPu compared to wildtype mice as measured by zero net flux with a concomitant decrease in evoked DA release and DA clearance measured by fast-scan cyclic voltammetry (FSCV).21 Our initial neurochemical analysis on CPu DA dynamics suggests that endogenous BDNF influences DA system homeostasis primarily by regulating release leading to adaptions in the DA uptake function.21 All of these studies suggest that BDNF can augment striatal DA functioning in a region-specific manner, but the mechanism of how BDNF modulates DA function still remains elusive. The present study uses FSCV to characterize the functional effects of TrkB agonists and inhibitors on DA dynamics in the CPu. By using striatal slices, it eliminates contributions from postsynaptic TrkB receptors allowing us to better understand how presynaptic TrkB receptors contribute to modulating DA dynamics. We hypothesized that a life-long reduction in BDNF would result in compensatory alterations in DA dynamics, which may be mediated by presynaptic TrkB activation. DA release and uptake have been evaluated in the striatum by using the endogenous ligand BDNF, the commercially available TrkB agonist 7,8-dihydroxyflavone (7,8-DHF), and the inhibitor K252a. This is the first reported study that examines the effect of TrkB agonist and an inhibitor on presynaptic DA dynamics using carbon fiber microelectrodes. While BDNF or a TrkB agonist increases DA release with no difference in DA clearance, a TrkB inhibitor decreases DA uptake with no effect on DA release. FSCV has the ability to assess the effect of non-DA receptors like the TrkB on striatal DA dynamics, and will help to provide a better understanding of how other systems contribute to the strength of synaptic DA transmission.