Hyun Yu

Macrophage migration inhibitory factor mediates the antidepressant actions of voluntary exercise

Voluntary exercise is known to have an antidepressant effect. However, the underlying mechanism for this antidepressant action of exercise remains unclear, and little progress has been made in identifying genes that are directly involved. We have identified macrophage migration inhibitory factor (MIF) by analyzing existing mRNA microarray data and confirmed the augmented expression of selected genes under two experimental conditions: voluntary exercise and electroconvulsive seizure. A proinflammatory cytokine, MIF is expressed in the central nervous system and involved in innate and adaptive immune responses. A recent study reported that MIF is involved in antidepressant-induced hippocampal neurogenesis, but the mechanism remains elusive. In our data, tryptophan hydroxylase 2 (Tph2) and brain-derived neurotrophic factor (Bdnf) expression were induced after MIF treatment in vitro, as well as during both exercise and electroconvulsive seizure in vivo. This increment of Tph2 was accompanied by increases in the levels of total serotonin in vitro. Moreover, the MIF receptor CD74 and the ERK1/2 pathway mediate the MIF-induced Tph2 and Bdnf gene expression as well as serotonin content. Experiments in Mif−/− mice revealed depression-like behaviors and a blunted antidepressant effect of exercise, as reflected by changes in Tph2 and Bdnf expression in the forced swim test. In addition, administration of recombinant MIF protein produced antidepressant-like behavior in rats in the forced swim test. Taken together, these results suggest a role of MIF in mediating the antidepressant action of exercise, probably by enhancing serotonin neurotransmission and neurotrophic factor-induced neurogenesis in the brain.

Repeated treatment with electroconvulsive seizures induces HDAC2 expression and down-regulation of NMDA receptor-related genes through histone deacetylation in the rat frontal cortex

The enzymatic activity of histone deacetylases (HDACs) leads to a histone deacetylation-mediated condensed chromatic structure, resulting in transcriptional repression, which has been implicated in the modifications of neural circuits and behaviors. Repeated treatment with electroconvulsive seizure (ECS) induces changes in histone acetylation, expression of various genes, and intrabrain cellular changes, including neurogenesis. In this study, we examined the effects of repeated ECS on the expression of class I HDACs and related changes in histone modifications and gene expression in the rat frontal cortex. Ten days of repeated ECS treatments (E10X) up-regulated HDAC2 expression at the mRNA and protein levels in the rat frontal cortex compared with sham-treated controls; this was evident in the nuclei of neuronal cells in the prefrontal, cingulate, orbital, and insular cortices. Among the known HDAC2 target genes, mRNA expression of N-methyl-d-aspartate (NMDA) receptor signaling-related genes, including early growth response-1 (Egr1), c-Fos, glutamate receptor, ionotropic, N-methyl d-aspartate 2A (Nr2a), Nr2b, neuritin1 (Nrn1), and calcium/calmodulin-dependent protein kinase II alpha (Camk2α), were decreased, and the histone acetylation of H3 and/or H4 proteins was also reduced by E10X. Chromatin immunoprecipitation analysis revealed that HDAC2 occupancy in the promoters of down-regulated genes was increased significantly. Moreover, administration of sodium butyrate, a HDAC inhibitor, during the course of E10X ameliorated the ECS-induced down-regulation of genes in the rat frontal cortex. These findings suggest that induction of HDAC2 by repeated ECS treatment could play an important role in the down-regulation of NMDA receptor signaling-related genes in the rat frontal cortex through histone modification.

The effect of clozapine on the AMPK-ACC-CPT1 pathway in the rat frontal cortex

Clozapine is an antipsychotic drug that has a greater efficacy than other medications in some contexts, especially for the treatment of treatment-resistant schizophrenia. However, clozapine induces more metabolic side-effects involving abnormality in lipid metabolism compared to other antipsychotics. AMP-activated protein kinase (AMPK) plays a central role in controlling lipid metabolism through modulating the downstream acetyl CoA carboxylase (ACC) and carnitine palmitoyl transferase 1 (CPT1) pathway. In this study, we investigated the effect of a single intraperitoneal injection of clozapine on the AMPK-ACC-CPT1 pathway in the rat frontal cortex, which has been implicated as a target site for this antipsychotic drug. At 2 h after injection, the clinically relevant dose of clozapine had activated AMPK, with increased phosphorylation of AMPKα at Thr(172), and had inactivated ACC, with increased phosphorylation of ACC at Ser(79). In addition, clozapine activated the brain-specific isoform of CPT1, CPT1c, whose activity is inhibited by unphosphorylated ACC, in the rat frontal cortex. Immunohistochemistry and immunofluorescence analysis showed that clozapine induced an increase in number of p-AMPKα (Thr(172))- and p-ACC (Ser(79))-positive cells among the neurons of the rat frontal cortex. Taken together, these results show that clozapine activated the AMPK-ACC-CPT1 pathway in the neurons of the rat frontal cortex. These findings indicate that the antipsychotic agent clozapine affects the lipid regulatory system of neurons in the brain.

Intracerebroventricular administration of ouabain, a Na/K-ATPase inhibitor, activates tyrosine hydroxylase through extracellular signal-regulated kinase in rat striatum

Alteration in dopamine neurotransmission has been reported to be involved in the mania of bipolar disorder. Tyrosine hydroxylase (TH) is the rate-limiting enzyme that is crucial for dopamine biosynthesis, and its activity is tightly regulated by phosphorylation at multiple N-terminal serine residues. Previously, we have reported that intracerebroventricular (ICV) injection of ouabain, a selective Na/K-ATPase inhibitor, induces hyperactivity in rats that mimics manic symptoms related to the activation of extracellular signal-regulated protein kinase1/2 (ERK1/2), which plays crucial roles in the modulation of TH phosphorylation. In this study, we investigated the effects of ICV injection of ouabain on TH phosphorylation in rat striatum and the involvement of ERK1/2 in ouabain-induced TH activation. ICV ouabain induced an acute dose-dependent increase in locomotor activity and in TH phosphorylation in rat striatum. TH phosphorylation at Ser19 was significantly increased with 100, 500, and 1000μM ouabain, and phosphorylation at Ser31 and Ser40 was significantly increased with 500 and 1000μM. We also found that ICV pretreatment with U0126, a specific MEK1/2 inhibitor, attenuated the 1000μM ouabain-induced increase in TH phosphorylation at Ser19, Ser31, and Ser40, as well as the hyperactivity of rats. Moreover, the increased phosphorylation of TH (Ser19, Ser31, and Ser40) was maintained until 8h after single administration ouabain was accompanied by increased phosphorylation of ERK1/2 (Thr202/Tyr204) and p90RSK (Thr359/Ser363). These findings imply that TH activation of the ERK1/2 signal pathway could play an important role in ouabain-induced hyperactivity of rats, a mania model.

A VANET routing based on the real-time road vehicle density in the city environment

The intelligent transportation system (ITS) can enhance the drivers safety by providing safety-related information such as traffic conditions and accident information to drivers. The vehicular ad hoc network (VANET) is an essential technology for the deployment of ITS. And, for the reliable delivery of safety-related information to vehicles in the VANET, a reliable VANET routing protocol is required. VANET routing is challenging since it is fundamentally different from conventional ad hoc networks; the vehicles move fast, and the network topology changes rapidly causing intermittent link connectivity. In this paper, we propose a routing protocol that works based on the real-time road vehicle density in order to provide fast and reliable communications so that it adapts to the dynamic vehicular city environment. In the proposed routing mechanism, each vehicle computes the vehicle density of the road to which it belongs by using beacon messages and the road information table. Based on the real-time road vehicle density information, each vehicle establishes a reliable route for packet delivery. In order to evaluate the performance of the proposed mechanism, we compare our proposed mechanism with GPSR through NS-2 based simulations and show that our mechanism outperforms GPSR in terms of delivery success rate and routing overhead.

Egr1 regulates lithium-induced transcription of the Period 2 (PER2) gene.

A growing body of evidence suggests that the circadian molecular system is involved in the pathogenic and therapeutic mechanisms underlying bipolar disorders. Lithium, a representative mood stabilizer, has been reported to induce the Period 2 (PER2) gene; however, the underlying molecular mechanisms require further study. We found that lithium upregulated PER2 expression at the transcriptional level in neuronally differentiated SH-SY5Y human neuroblastoma cells. Promoter reporter analyses using serial deletions of the PER2 promoter revealed that two early growth response 1 (Egr1)-binding sites (EBS) between positions -180 and -100 are required for maximal activation of the PER2 promoter by lithium. Ectopic expression of Egr1 enhanced lithium-induced PER2 promoter activity, while a point mutation in EBS abolished it. Electrophoretic mobility shift assays and chromatin immunoprecipitation indicated that Egr1 bound directly to the PER2 promoter. Stimulation of the extracellular-signal regulated kinase (ERK)1/2/Elk1 pathway by lithium was functionally linked to PER2 expression through Egr1 induction, and lithium-induced PER2 expression was strongly attenuated by depletion of Egr1 by siRNA. Lithium also upregulated the expression of Per2 and Egr1 in mouse frontal cortex. Induction of Per2 by lithium was attenuated in Egr1(-/-) mice. In conclusion, lithium stimulates PER2 transcription through the ERK/Elk1/Egr1 pathway in neuronal cells, indicating a connection between the ERK-Egr1 pathway and a circadian gene system in the mechanism of action of lithium.

The effect of cyclosporine A on the phosphorylation of the AMPK pathway in the rat hippocampus.

Cyclosporine A (CsA), an immunosuppressant and calcineurin inhibitor, induces hyperlipidemia in humans and animals. AMP-activated protein kinase (AMPK) is involved in metabolic homeostasis and lipid metabolism through modulating downstream molecules acetyl CoA carboxylase (ACC) and 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoAR). AMPK activity is regulated by the phosphorylation at the Thr-172 residue by its upstream liver kinase B 1 (LKB1), Ca(2+)/calmodulin-dependent protein kinase kinase β (CaMKKβ) or transforming growth-factor-β-activated kinase 1 (TAK1). AMPK can be deactivated through dephosphorylation by protein phosphatase 2Cα (PP2Cα). In this study, we demonstrated that phosphorylation at Thr-172-AMPK increased with a concurrent increase in the phosphorylation of Ser-431-LKB1 and Thr-184/187-TAK1 in the rat hippocampus at 5 h after an intraperitoneal CsA (50 mg/kg) injection. CsA did not affect the phosphorylation of Thr-196-Ca(2+)/calmodulin-dependent protein kinase 4 (CaMK4) and the amount of PP2Cα. An increased phosphorylation of Ser-79-ACC and Ser-872-HMG-CoAR was also observed. In conclusion, our data indicate that CsA activates the AMPK pathway in the rat hippocampus, which suggests that CsA affects the regulatory signaling pathway of lipid metabolism in the rat brain.

Electroconvulsive seizure increases phosphorylation of PKC substrates, including GAP-43, MARCKS, and neurogranin, in rat brain

Protein kinase C (PKC) has been suggested as a molecular target related to the pathogenetic and therapeutic mechanisms of mood disorders in which electroconvulsive seizure (ECS) is effective. However, the reports concerning the effects of ECS on PKC are anecdotal and need further clarification. In this study, we examined the effects of ECS treatment on the phosphorylation of PKC substrates, including GAP-43, MARCKS, and neurogranin. Immunoblot using anti-p-PKC substrate antibodies revealed that a single ECS treatment induced temporal changes in the phosphorylation level of PKC substrates in rat brain, reflecting the effects on PKC activity. Phosphorylation of GAP-43 and MARCKS, representative PKC substrates related to synaptic remodeling, increased from 5 to 30 min, after a transient decrease at 0 min immediately after ECS, and returned to basal levels at 60 min in rat frontal cortex, hippocampus, and cerebellum. Phosphorylation of neurogranin, another PKC substrate, showed a similar pattern of temporal changes in the frontal cortex and hippocampus. Immunohistochemical analysis revealed that p-GAP-43 and p-MARCKS were densely stained throughout the neuronal cells of the prefrontal cortex and hippocampus, and the Purkinje cells of cerebellum, after ECS treatment. Brief and transient activation of PKC may be translated into long-term biochemical changes, resulting in synaptic plasticity. Taken together, the acute effects of ECS on PKC activity, which could be an underpinning of long-term biochemical changes induced by ECS, may contribute to understand the molecular mechanism of ECS.

A Stable Routing Protocol for Vehicles in Urban Environments

The vehicular ad hoc network (VANET) is an essential technology that enables the deployment of the intelligent transportation system (ITS), which improves the traffic safety and efficiency. For the efficient message delivery in VANETs, it is desirable to provide a reliable and stable VANET routing protocol. However, VANET routing is challenging since the VANET is fundamentally different from conventional wireless ad hoc networks; vehicles move fast, and the network topology changes rapidly, causing intermittent and dynamic link connectivity. In this paper, we propose a VANET routing protocol that works based on the real-time road vehicle density information in order to provide fast and reliable message delivery so that it can adapt to the dynamic vehicular urban environment. In the proposed mechanism, each vehicle computes the real-time traffic density of the road to which it belongs from the beacon messages sent by vehicles on the opposite lane and its road information table. Using the road traffic density information as a routing metric, each vehicle establishes a reliable route for packet delivery. We compare our proposed mechanism with the well-known GPSR via NS-2 based simulations and show that our mechanism outperforms GPSR in terms of both delivery success rate and routing overhead.

Node Movement Detection to Overcome False Route Failures in Mobile Ad Hoc Networks

The mobile ad hoc network (MANET) is a wireless network without the wired infrastructure such as base stations in which mobile nodes communicate via multiple wireless links. In the MANET, route failures may happen because of node movement or wireless link collisions on routes. Since route failures due to wireless link collisions (i.e., false route failures) are not from network topology changes, they should not trigger route reestablishments; otherwise, the network performance will be aggravated. In this paper, we propose a node movement detection scheme that can prevent unnecessary route reestablishments caused by false route failures in the MANET. The proposed mechanism allows a node to decide its movement based on the reception status of modified HELLO messages which indicates the degree of its local topology changes. By carrying out NS-2 based simulations, we show that our node movement detection scheme resolves the false route failure problem more effectively than the existing mechanisms.