Hiroyuki Nawa

Activation of the TrkB Neurotrophin Receptor Is Induced by Antidepressant Drugs and Is Required for Antidepressant-Induced Behavioral Effects

Recent studies have indicated that exogenously administered neurotrophins produce antidepressant-like behavioral effects. We have here investigated the role of endogenous brain-derived neurotrophic factor (BDNF) and its receptor trkB in the mechanism of action of antidepressant drugs. We found that trkB.T1-overexpressing transgenic mice, which show reduced trkB activation in brain, as well as heterozygous BDNF null (BDNF+/−) mice, were resistant to the effects of antidepressants in the forced swim test, indicating that normal trkB signaling is required for the behavioral effects typically produced by antidepressants. In contrast, neurotrophin-3+/− mice showed a normal behavioral response to antidepressants. Furthermore, acute as well as chronic antidepressant treatment induced autophosphorylation and activation of trkB in cerebral cortex, particularly in the prefrontal and anterior cingulate cortex and hippocampus. Tyrosines in the trkB autophosphorylation site were phosphorylated in response to antidepressants, but phosphorylation of the shc binding site was not observed. Nevertheless, phosphorylation of cAMP response element-binding protein was increased by antidepressants in the prefrontal cortex concomitantly with trkB phosphorylation and this response was reduced in trkB.T1-overexpressing mice. Our data suggest that antidepressants acutely increase trkB signaling in a BDNF-dependent manner in cerebral cortex and that this signaling is required for the behavioral effects typical of antidepressant drugs. Neurotrophin signaling increased by antidepressants may induce formation and stabilization of synaptic connectivity, which gradually leads to the clinical antidepressive effects and mood recovery.

Brain-derived neurotrophic factor induces mammalian target of rapamycin-dependent local activation of translation machinery and protein synthesis in neuronal dendrites.

In neurons, perisynaptic or dendritic translation is implicated in synapse-wide alterations of function and morphology triggered by neural activity. The molecular mechanisms controlling local translation activation, however, have yet to be elucidated. Here, we show that local protein synthesis and translational activation in neuronal dendrites are upregulated by brain-derived neurotrophic factor (BDNF) in a rapamycin and small interfering RNA specific for mammalian target of rapamycin (mTOR)-sensitive manner. In parallel, BDNF induced the phosphorylation of tuberin and the activation of mTOR in dendrites and the synaptoneurosome fraction. mTOR activation stimulated translation initiation processes involving both eIF4E/4E-binding protein (4EBP) and p70S6 kinase/ribosomal S6 protein. BDNF induced phosphorylation of 4EBP in isolated dendrites. Moreover, local puff application of BDNF to dendrites triggered S6 phosphorylation in a restricted area. Taken together, these data indicate that mTOR-dependent translation activation is essential for the upregulation of local protein synthesis in neuronal dendrites.

Abnormal expression of brain-derived neurotrophic factor and its receptor in the corticolimbic system of schizophrenic patients.

Previous neuropathological studies have revealed that the corticolimbic system of schizophrenic patients expresses abnormal levels of various synaptic molecules, which are known to be influenced by the neuronal differentiation factors, neurotrophins. Therefore, we determined levels of neurotrophins and their receptors in the postmortem brains of schizophrenic patients and control subjects in relation to molecular impairments in schizophrenia. Among the neurotrophins examined, levels of brain-derived neurotrophic factor (BDNF) were elevated specifically in the anterior cingulate cortex and hippocampus of schizophrenic patients, but levels of nerve growth factors and neurotrophin-3 showed no change in any of the regions examined. In parallel, the expressions of TrkB receptor and calbindin-D, which are both influenced by BDNF, were reduced significantly in the hippocampus or the prefrontal cortex. However, neuroleptic treatment did not appear to mimic the neurotrophic change. Neither withdrawal of drug treatment in patients nor chronic administration of haloperidol to rats altered levels of BDNF. These findings suggest that neurotrophic abnormality is associated with the corticolimbic structures of schizophrenic patients and might provide the molecular substrate for pathological manifestations of the illness.

Cytokine and growth factor involvement in schizophrenia—support for the developmental model

Medical treatment with various cytokines can provoke psychiatric symptoms. Conversely, psychiatric patients can display abnormalities in cytokine and neurotrophic factor expression. Such observations have pointed to the potential contribution of cytokines and growth factors to schizophrenic pathology and/or etiology. The cellular targets of the relevant factors and the nature of their actions remain to be explored in mental illness, however. Recent physiological studies demonstrate that cytokines and neurotrophic factors can markedly influence synaptic transmission and plasticity upon acute or chronic application. Moreover, many of the molecular alterations observed in the schizophrenic brain are consistent with abnormalities in cytokine and neurotrophic factor regulation of these molecules. In this review, we summarize these molecular pathology findings for schizophrenia and highlight the neurodevelopmental activities of cytokines and neurotrophic factors that may contribute to the etiology or pathology of this illness.

Phosphatidylinositol 3-kinase: a molecule mediating BDNF-dependent spatial memory formation

Brain-derived neurotrophic factor (BDNF) plays a critical role in synaptic plasticity such as long-term potentiation (LTP), a form of synaptic correlate of learning and memory. BDNF is also implicated in learning and memory. We have demonstrated that radial arm maze training in rats for spatial learning and memory results in a significant increase in the BDNF mRNA expression in the hippocampus. Moreover, antisense BDNF oligonucleotide treatment impaired not only acquisition, but also maintenance and/or recall of spatial memory in the maze. Although these results suggest a role of BDNF for spatial memory processes, the signal transduction mechanisms that mediate the actions of BDNF remain unknown. Here we show that phosphorylation of BDNF receptor tyrosine kinase B (TrkB), phosphatidylinositol 3-kinase (PI3-K) and Akt, a target of PI3-K, in the hippocampus increased in parallel with spatial memory formation. Moreover, an activation of translational processes was suggested in the hippocampus after the maze training. When spatial learning was inhibited by antisense BDNF oligodeoxynucleotide, the activation was diminished. Chronic treatment with PI3-K inhibitor wortmannin impaired spatial learning. Our findings suggested that activation of TrkB/PI3-K and protein synthesis signaling pathway by BDNF in the hippocampus is important for spatial memory.

Brain-derived neurotrophic factor regulates the expression of AMPA receptor proteins in neocortical neurons.

The role of the neurotrophins; nerve growth factor, brain-derived neurotrophic factor, neurotrophin-3 and neurotrophin-4/5, in synaptic development and plasticity has been extensively investigated. The neurotrophins regulate synaptic transmission as well as neural development in the brain. However, the mechanisms underlying these processes are unknown. In this study we show that brain-derived neurotrophic factor triggers an increase in alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptor (GluR) proteins without significant changes in their messenger RNA levels. Brain-derived neurotrophic factor treatment specifically increased the protein levels of GluR1 (193+/-22%) and GluR2/3 (182+/-11%) in cultured rat neocortical neurons. In contrast, nerve growth factor and neurotrophin-3 failed to alter the protein levels of these neurons, and brain-derived neurotrophic factor effects on N-methyl-D-aspartate-type glutamate receptors were either modest or negligible. Immunocytochemical studies indicated that the increase in AMPA receptor proteins reflects the induction of their neuronal expression, but not selective neuronal survival. In agreement with these results, cortical neurons from brain-derived neurotrophic factor-knockout mice exhibited a reduction in AMPA receptor proteins in the cytoskeletal fraction containing postsynaptic proteins. Thus, the neurotrophin plays a crucial role in modulating the expression of AMPA receptors presumably at translational or post-translation levels and is implicated in synaptic development and plasticity.

Biological characterization and optical imaging of brain-derived neurotrophic factor-green fluorescent protein suggest an activity-dependent local release of brain-derived neurotrophic factor in neurites of cultured hippocampal neurons.

To visualize the release dynamics of the brain‐derived neurotrophic factor (BDNF) involved in neural plasticity, we constructed a plasmid encoding green fluorescent protein (GFP) fused with BDNF. First, several biological studies confirmed that this fusion protein (BDNF‐GFP) mimics the biological functions and the release kinetics of unfused (native) BDNF. Second, when BDNF‐GFP was expressed in cultured hippocampal neurons, we observed that this protein formed striking clusters in the neurites of mature neurons and colocalized with the PSD‐95 immunoreactivity. Such a clustered BDNF‐GFP rapidly disappeared in response to depolarization with KCl, as revealed by confocal microscopic studies. These data suggest that BDNF is locally and rapidly released at synaptic sites in an activity‐dependent manner. Optical studies using BDNF‐GFP may provide important evidence regarding the participation of BDNF in synaptic plasticity. J. Neurosci. Res. 64:1–10, 2001.

Abnormal expression of epidermal growth factor and its receptor in the forebrain and serum of schizophrenic patients

Epidermal growth factor (EGF) comprises a structurally related family of proteins containing heparin-binding EGF-like growth factor (HB-EGF) and transforming growth factor alpha (TGFα) that regulates the development of dopaminergic neurons as well as monoamine metabolism. We assessed the contribution of EGF to schizophrenia by measuring EGF family protein levels in postmortem brains and in fresh serum of schizophrenic patients and control subjects. EGF protein levels were decreased in the prefrontal cortex and striatum of schizophrenic patients, whereas the levels of HB-EGF and TGFα were not significantly different in any of the regions examined. Conversely, EGF receptor expression was elevated in the prefrontal cortex. Serum EGF levels were markedly reduced in schizophrenic patients, even in young, drug-free patients. Chronic treatment of animals with the antipsychotic drug haloperidol had no influence on EGF levels in the brain or serum. These findings suggest that there is abnormal EGF production in various central and peripheral tissues of patients with both acute and chronic schizophrenia. EGF might thus provide a molecular substrate for the pathologic manifestation of the illness, although additional studies are required to determine a potential link between impaired EGF signaling and the pathology/etiology of schizophrenia.

Prefrontal Abnormality of Schizophrenia Revealed by DNA Microarray: Impact on Glial and Neurotrophic Gene Expression

Abstract: DNA microarrays with isotope labeling from gene‐specific primers enable sensitive detection of rare mRNAs, including neurotrophin and cytokine mRNAs in the brain. Using high‐quality RNA from postmortem brains, gene‐expression profiles covering 1373 genes were assessed in the dorsoprefrontal cortex of schizophrenic patients and compared with those of nonpsychiatric subjects. Statistical analysis of the DNA microarray data confirmed the findings of a previous GeneChip study by Hakak et al. (Proc. Natl. Acad. Sci. USA Vol. 98, pp. 4746‐4751, 2001). The highest frequency of mRNA expression alterations occurred in oligodendrocyte‐ and astrocyte‐related genes in the prefrontal cortex of schizophrenic patients, followed by the category for the genes for growth factors/neurotrophic factors and their receptors. Whether each mRNA signal represents the expression of the individual genes or homologous genes in the category remains to be determined, however. To control for potential medication effects on patients, RNA from cynomolgus monkeys that were treated with haloperidol for 3 months was also subjected to DNA microarray analysis. A few genes overlapped between the gene‐expression profiles of the monkeys and patients. The present profiling study suggests a potential biological link between abnormal neurotrophic signals and impaired glial functions in schizophrenic pathology.

Biochemical evidence for localization of AMPA-type glutamate receptor subunits in the dendritic raft

A low density Triton-insoluble fraction with characteristic lipid composition was prepared from synaptic plasma membrane from the rat forebrain. The fraction was named dendritic raft based on its absence of the presynaptic marker synaptophysin, the presence of postsynaptic Glutamate receptor (GluR) subunits, and its resemblance to raft, caveolae-like structure. We found a differential distribution of NMDA-type and AMPA-type GluR subunits in the dendritic raft and postsynaptic density (PSD) fractions; the latter type GluR subunits were localized to the dendritic raft as well as PSD fraction, whereas the former type was mostly localized to the PSD fraction. We also found the differential distribution of the components of ras/mitogen-activated protein kinase (MAPK) pathway to the dendritic raft and PSD fractions. Dendritic raft and PSD may possibly interact at the postsynaptic sites for efficient signal processing that is required for expression of synaptic plasticity.