Extracellular factors may act on cells in two distinct modes: an acute increase in concentration due to regulated secretion, or a gradual increase in concentration when secreted constitutively or from a distant source. We show that cellular responses to BDNF differ dramatically depending on how BDNF is delivered. In cultured neurons, acute and gradual increases in BDNF elicited, respectively, transient and sustained activation of TrkB receptor and its downstream signaling, leading to differential expression of Homer1a and Arc. Transient TrkB activation promoted neurite elongation and spine head enlargement, whereas sustained TrkB activation facilitated neurite branch and spine neck elongation. In hippocampal slices, fast and slow increases in BDNF enhanced basal synaptic transmission and LTP, respectively. Thus, the kinetics of TrkB activation is critical for cell signaling and functions. This temporal dimension in cellular signaling may also have implications for the therapeutic drug design.

Organized neuronal firing is critical for cortical processing and is disrupted in schizophrenia. Using 5’ RACE in human brain, we identified a primate-specific isoform (3.1) of the K+-channel KCNH2 that modulates neuronal firing. KCNH2-3.1 mRNA levels are comparable to KCNH2-1A in brain, but 1000-fold lower in heart. In schizophrenic hippocampus, KCNH2-3.1 expression is 2.5-fold greater than KCNH2-1A. A meta-analysis of 5 clinical samples (367 families, 1158 unrelated cases, 1704 controls) shows association of SNPs in KCNH2 with schizophrenia. Risk-associated alleles predict lower IQ scores and speed of cognitive processing, altered memory-linked fMRI signals, and increased KCNH2-3.1 expression in post-mortem hippocampus. KCNH2-3.1 lacks a domain critical for slow channel deactivation. Overexpression of KCNH2-3.1 in primary cortical neurons induces a rapidly deactivating K+ current and a high-frequency, non-adapting firing pattern. These results identify a novel KCNH2 channel involved in cortical physiology, cognition, and psychosis, providing a potential new psychotherapeutic drug target.

Dysbindin-1 regulates D2-receptor trafficking and is implicated in schizophrenia and related cognitive abnormalities, but whether this molecular effect mediates the clinical manifestations of the disorder is unknown. We explored in dysbindin-1-deficient mice (dys–/–) (1) schizophrenia-related behaviors, (2) molecular and electrophysiological changes in medial prefrontal cortex (mPFC) and (3) the dependence of these on D2-receptor stimulation. Dysbindin-1 disruption altered dopamine-related behaviors and impaired working memory under challenging/stressful conditions. Dys–/– pyramidal neurons in mPFC layers II/III were hyperexcitable at baseline but hypoexcitable following D2 stimulation. Dys–/– were also respectively more and less sensitive to D2 agonist- and antagonist-induced behavioral effects. Dys–/– had reduced expression of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and CaMKKβ in mPFC. Chronic D2 agonist treatment reproduced these changes in protein expression, and some of the dys–/– behavioral effects. These results elucidate dysbindin's modulation of D2-related behavior, cortical activity and mPFC CaMK components, implicating cellular and molecular mechanisms of the association of dysbindin with psychosis.

Dysbindin has been implicated in the pathogenesis of schizophrenia, but little is known about how dysbindin affects neuronal function in the circuitry underlying psychosis and related behaviors. Using a dysbindin knockout line (dys ؊/؊ ) derived from the natural dysbindin mutant Sandy mice, we have explored the role of dysbindin in dopamine signaling and neuronal function in the prefrontal cortex (PFC). Combined cell imaging and biochemical experiments revealed a robust increase in the dopamine receptor D2, but not D1, on cell surface of neurons from dys ؊/؊ cortex. This was due to an enhanced recycling and insertion, rather than reduced endocytosis, of D2. Disruption of dysbindin gene resulted in a marked decrease in the excitability of fast-spiking (FS) GABAergic interneurons in both PFC and striatum. Dys ؊/؊ mice also exhibited a decreased inhibitory input to pyramidal neurons in layer V of PFC. The increased D2 signaling in dys ؊/؊ FS interneurons was associated with a more pronounced increase in neuronal firing in response to D2 agonist, compared to that in wild-type interneurons. Taken together, these results suggest that dysbindin regulates PFC function by facilitating D2-mediated modulation of GABAergic function.dopamine D2 receptor ͉ schizophrenia ͉ prefrontal cortex G enetic variants in a gene on 6p22.3, dysbindin (DTNBP1), have been shown to be one of the several genes that are associated with schizophrenia (1). Schizophrenia patients have significantly reduced expression of dysbindin mRNA and protein in prefrontal cortex and hippocampus (2, 3). While it remains unclear how changes in dysbindin expression could contribute to the pathogenesis of schizophrenia, cell biological studies have begun to address the physiological function of dysbindin in neurons. Downregulation of dysbindin by siRNA in cultured neurons leads to decreases in the expression of SNAP25 and levels of extracellular glutamate or dopamine (4, 5). Dysbindin contributes to normal biogenesis of lysosome-related organelles (LROs) by binding to proteins in the BLOC-1 complex (6, 7), which regulates trafficking of LROs. The Sandy mouse (Sdy), which harbors an in-frame deletion of two exons of the dysbindin gene (8), exhibits a reduced readily releasable pool of synaptic vesicles and larger vesicle size (9). Although dysbindin protein is localized both pre-and postsynaptically (7), little is known about its postsynaptic function. Recently, downregulation of dysbindin has been shown to increase cell surface expression of dopamine receptor D2 (D2), but not dopamine receptor D1 (D1), in human SH-SY5Y neuroblastoma cells and in cultured cortical neurons (10). Dopamine receptor internalization (or endocytosis) is a general mechanism to adjust neuronal responses to dopamine stimulation. Both D1 and D2 are G protein coupled receptors (GPCRs) that undergo constitutive and ligand-induced internalization. Unlike D1, which is recycled back to the plasma membrane after endocytosis, D2 is generally trafficked to the lysosomal pathway and degraded (11)(1...

Brain-derived neurotrophic factor (BDNF) has been shown to regulate neuronal survival and synaptic plasticity in the central nervous system (CNS) in an activity-dependent manner, but the underlying mechanisms remain unclear. Here we report that the number of BDNF receptor TrkB on the surface of hippocampal neurons can be enhanced by high frequency neuronal activity and synaptic transmission, and this effect is mediated by Ca2+ influx. Using membrane protein biotinylation as well as receptor binding assays, we show that field electric stimulation increased the number of TrkB on the surface of cultured hippocampal neurons. Immunofluorescence staining suggests that the electric stimulation facilitated the movement of TrkB from intracellular pool to the cell surface, particularly on neuronal processes. The number of surface TrkB was regulated only by high frequency tetanic stimulation, but not by low frequency stimulation. The activity dependent modulation appears to require Ca2+ influx, since treatment of the neurons with blockers of voltage-gated Ca2+ channels or NMDA receptors, or removal of extracellular Ca2+, severely attenuated the effect of electric stimulation. Moreover, inhibition of Ca2+/calmodulin-dependent kinase II (CaMKII) significantly reduced the effectiveness of the tetanic stimulation. These findings may help us to understand the role of neuronal activity in neurotrophin function and the mechanism for receptor tyrosine kinase signaling.

Formation of specific neuronal connections often involves competition between adjacent axons, leading to stabilization of the active terminal, while retraction of the less active ones. The underlying molecular mechanisms remain unknown. We show that activitydependent conversion of pro-brain-derived neurotrophic factor (proBDNF) to mature (m)BDNF mediates synaptic competition. Stimulation of motoneurons triggers proteolytic conversion of proBDNF to mBDNF at nerve terminals. In Xenopus nerve-muscle cocultures, in which two motoneurons innervate one myocyte, proBDNFp75 NTR signaling promotes retraction of the less active terminal, whereas mBDNF-tyrosine-related kinase B (TrkB) p75NTR (p75 neurotrophin receptor) facilitates stabilization of the active one. Thus, proBDNF and mBDNF may serve as potential "punishment" and "reward" signals for inactive and active terminals, respectively, and activity-dependent conversion of proBDNF to mBDNF may regulate synapse elimination. neuromuscular junction | pro-neurotrophin | synapse competition

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Signaling mechanisms underlying neurotrophic regulation of synaptic transmission are not fully understood. Here we show that neurotrophin-3 (NT3)-induced potentiation of synaptic transmission at the neuromuscular synapses is blocked by inhibition of phosphoinositide-3 kinase, phospholipase C-gamma or the downstream IP3 receptors of phospholipase C-gamma, but not by inhibition of MAP kinase. However, neither stimulation of Ca2+ release from intracellular stores by photolysis of caged IP3, nor expression of a constitutively active phosphoinositide-3 kinase (PI3K*) in presynaptic motoneurons alone is sufficient to enhance transmission. Photo-uncaging of IP3 in neurons expressing PI3K* elicits a marked synaptic potentiation, mimicking the NT3 effect. These results reveal an involvement of PI3 kinase in transmitter release, and suggest that concomitant activation of PI3 kinase and IP3 receptors is both necessary and sufficient to mediate the NT3-induced synaptic potentiation.