Biomedical subjects

Rami Yaka

Publications and source records attributed to Rami Yaka.

11 recordsLinked to original sources

PubMed2006

Cocaine enhances NMDA receptor-mediated currents in ventral tegmental area cells via dopamine D5 receptor-dependent redistribution of NMDA receptors.

Cocaine-induced plasticity of glutamatergic synaptic transmission in the ventral tegmental area (VTA) plays an important role in brain adaptations that promote addictive behaviors. However, the mechanisms responsible for triggering these synaptic changes are unknown. Here, we examined the effects of acute cocaine application on glutamatergic synaptic transmission in rat midbrain slices. Cocaine caused a delayed increase in NMDA receptor (NMDAR)-mediated synaptic currents in putative VTA dopamine (DA) cells. This effect was mimicked by a specific DA reuptake inhibitor and by a DA D1/D5 receptor agonist. The effect of cocaine was blocked by a DA D1/D5 receptor antagonist as well as by inhibitors of the cAMP/cAMP-dependent protein kinase A (PKA) pathway. Furthermore, biochemical analysis showed an increase in the immunoreactivity of the NMDAR subunits NR1 and NR2B and their redistribution to the synaptic membranes in VTA neurons. Accordingly, NMDAR-mediated EPSC decay time kinetics were significantly slower after cocaine, suggesting an increased number of NR2B-containing NMDARs. Finally, pharmacological analysis indicates that NR2B subunits might be incorporated in triheteromeric NR1/NR2A/NR2B complexes rather than in "pure" NR1/NR2B NMDA receptors. Together, our data suggest that acute cocaine increases NMDAR function in the VTA via activation of the cAMP/PKA pathway mediated by a DA D5-like receptor, leading to the insertion of NR2B-containing NMDARs in the membrane. These results provide a potential mechanism by which acute cocaine promotes synaptic plasticity of VTA neurons, which could ultimately lead to the development of addictive behaviors.

Animals↗

PubMed2004

Spatial and temporal regulation of RACK1 function and N-methyl-D-aspartate receptor activity through WD40 motif-mediated dimerization.

Efficient signaling requires accurate spatial and temporal compartmentalization of proteins. RACK1 is a scaffolding protein that fulfils this role through interaction of binding partners with one of its seven WD40 domains. We recently identified the kinase Fyn and the NR2B subunit of the N-methyl-D-Aspartate receptor (NMDAR) as binding partners of RACK1. Scaffolding of Fyn near its substrate NR2B by RACK1 inhibits Fyn phosphorylation of NR2B and thereby negatively regulates channel function. We found that Fyn and NR2B share the same binding site on RACK1; however, their binding to RACK1 is not mutually exclusive (Yaka, R., Thornton, C., Vagts, A. J., Phamluong, K., Bonci, A., and Ron, D. (2002) Proc. Natl. Acad. Sci. U. S. A. 99, 5710-5715). We therefore tested the hypothesis that RACK1 forms a homodimer that allows the simultaneous binding of Fyn and NR2B. We found that RACK1 binds to itself both in vitro and in the brain. Deletion analyses identified a RACK1-RACK1 dimer-binding site within the 4th WD40 repeat, and application of the 4th WD40 repeat or a peptide derivative to hippocampal slices inhibited NMDAR activity. We further found that in hippocampal slices, both RACK1 and NR2B associated with another WD40 protein, the beta-subunit of G protein (Gbeta), previously shown to heterodimerize with RACK1 in vitro (Dell, E. J., Connor, J., Chen, S., Stebbins, E. G., Skiba, N. P., Mochly-Rosen, D., and Hamm, H. E. (2002) J. Biol. Chem. 277, 49888-49895). However, activation of the pituitary adenylate cyclase polypeptide (1-38) G protein-coupled receptor, previously found to induce the dissociation of RACK1 from the NMDAR complex (Yaka, R., He, D. Y., Phamluong, K., and Ron, D. (2003) J. Biol. Chem. 278, 9630-9638), attenuated the association of Gbeta with RACK1 and NR2B. Based on these results, we propose that WD40-mediated homo- and heterodimerization of RACK1 mediate the formation of a transient signaling complex that includes the NMDAR, a G protein and Fyn.

Amino Acid Sequence↗

PubMed2003

Corticotropin-releasing factor requires CRF binding protein to potentiate NMDA receptors via CRF receptor 2 in dopamine neurons.

Stress increases addictive behaviors and is a common cause of relapse. Corticotropin-releasing factor (CRF) plays a key role in the modulation of drug taking by stress. However, the mechanism by which CRF modulates neuronal activity in circuits involved in drug addiction is poorly understood. Here we show that CRF induces a potentiation of NMDAR (N-methyl-D-aspartate receptor)-mediated synaptic transmission in dopamine neurons of the ventral tegmental area (VTA). This effect involves CRF receptor 2 (CRF-R2) and activation of the phospholipase C (PLC)-protein kinase C (PKC) pathway. We also find that this potentiation requires CRF binding protein (CRF-BP). Accordingly, CRF-like peptides, which do not bind the CRF-BP with high affinity, do not potentiate NMDARs. These results provide evidence of the first specific roles for CRF-R2 and CRF-BP in the modulation of neuronal activity and suggest that NMDARs in the VTA may be a target for both drugs of abuse and stress.

Animals↗

PubMed2003

Scaffolding of Fyn kinase to the NMDA receptor determines brain region sensitivity to ethanol.

Alcohol (ethanol) abuse is a major societal problem. Although ethanol is a structurally simple, diffusible molecule, its sites of action are surprisingly selective, and the molecular mechanisms underlying specificity in ethanol actions are not understood. The NMDA receptor channel is one of the main targets for ethanol in the brain. We report here that the brain region-specific compartmentalization of Fyn kinase determines NMDA receptor sensitivity to ethanol. We demonstrate that, in the hippocampus but not in the cerebral cortex, Fyn is targeted to the NR2B subunit of the NMDA receptor by the scaffolding protein RACK1. During acute exposure to ethanol, RACK1 is dissociated from the complex, thereby facilitating Fyn-mediated phosphorylation of NR2B, which enhances channel activity, counteracting the inhibitory actions of ethanol. In this way, the selective scaffolding can account for the ethanol-induced acute tolerance of NMDA receptor activity that is detected in the hippocampus but not in the cerebral cortex. The phosphorylation-dependent, region-specific activities of ethanol on the NMDA receptor provide a compelling molecular explanation that accounts for the selective activities of ethanol and may have important implications for elucidating pathways leading to alcohol addiction.

Animals↗

PubMed2003

H-Ras modulates N-methyl-D-aspartate receptor function via inhibition of Src tyrosine kinase activity.

Tyrosine phosphorylation of the NR2A and NR2B subunits of the N-methyl-d-aspartate (NMDA) receptor by Src protein-tyrosine kinases modulates receptor channel activity and is necessary for the induction of long term potentiation (LTP). Deletion of H-Ras increases both NR2 tyrosine phosphorylation and NMDA receptor-mediated hippocampal LTP. Here we investigated whether H-Ras regulates phosphorylation and function of the NMDA receptor via Src family protein-tyrosine kinases. We identified Src as a novel H-Ras binding partner. H-Ras bound to Src but not Fyn both in vitro and in brain via the Src kinase domain. Cotransfection of H-Ras and Src inhibited Src activity and decreased NR2A tyrosine phosphorylation. Treatment of rat brain slices with Tat-H-Ras depleted NR2A from the synaptic membrane, decreased endogenous Src activity and NR2A phosphorylation, and decreased the magnitude of hippocampal LTP. No change was observed for NR2B. We suggest that H-Ras negatively regulates Src phosphorylation of NR2A and retention of NR2A into the synaptic membrane leading to inhibition of NMDA receptor function. This mechanism is specific for Src and NR2A and has implications for studies in which regulation of NMDA receptor-mediated LTP is important, such as synaptic plasticity, learning, and memory and addiction.

Animals↗

PubMed2003

Pituitary adenylate cyclase-activating polypeptide (PACAP(1-38)) enhances N-methyl-D-aspartate receptor function and brain-derived neurotrophic factor expression via RACK1.

We recently identified a novel mechanism for modulation of the phosphorylation state and function of the N-methyl-d-aspartate (NMDA) receptor via the scaffolding protein RACK1. We found that RACK1 binds both the NR2B subunit of the NMDA receptor and the nonreceptor protein-tyrosine kinase, Fyn. RACK1 inhibits Fyn phosphorylation of NR2B and decreases NMDA receptor-mediated currents in CA1 hippocampal slices (Yaka, R., Thornton, C., Vagts, A. J., Phamluong, K., Bonci, A., and Ron, D. (2002) Proc. Natl. Acad. Sci. U. S. A. 99, 5710-5715). Here, we identified the signaling cascade by which RACK1 is released from the NMDA receptor complex and identified the consequences of the dissociation. We found that activation of the cAMP/protein kinase A pathway in hippocampal slices induced the release of RACK1 from NR2B and Fyn. This resulted in the induction of NR2B phosphorylation and the enhancement of NMDA receptor-mediated activity via Fyn. We identified the neuropeptide, pituitary adenylate cyclase activating polypeptide (PACAP(1-38)), as a ligand that induced phosphorylation of NR2B and enhanced NMDA receptor potentials. Finally, we found that activation of the cAMP/protein kinase A pathway induced the movement of RACK1 to the nuclear compartment in dissociated hippocampal neurons. Nuclear RACK1 in turn was found to regulate the expression of brain-derived neurotrophic factor induced by PACAP(1-38). Taken together our results suggest that activation of adenylate cyclase by PACAP(1-38) results in the release of RACK1 from the NMDA receptor and Fyn. This in turn leads to NMDA receptor phosphorylation, enhanced activity mediated by Fyn, and to the induction of brain-derived neurotrophic factor expression by RACK1.

Adenylyl Cyclases↗

PubMed2003

Cellular adaptation to chronic ethanol results in altered compartmentalization and function of the scaffolding protein RACK1.

BACKGROUND: Previously, we found that acute ethanol induces the translocation of the scaffolding protein RACK1 to the nucleus. Recently, we found that nuclear RACK1 mediates acute ethanol induction of immediate early gene c-fos expression. Alterations in gene expression are thought to lead to long-term changes that ultimately contribute to the development of alcohol addiction and toxicity. Therefore, we sought to determine the effects of chronic exposure of cells to ethanol on the cellular compartmentalization of RACK1 and on c-fos messenger RNA (mRNA) and protein expression. METHODS: Rat C6 glioma cells were used as the cell culture model. Immunohistochemistry was implemented to visualize the localization of RACK1 and to monitor the protein level of c-fos. Reverse-transcription polymerase chain reaction was used to measure c-fos mRNA levels. The Tat-protein transduction method was used to transduce recombinant Tat-RACK1 into cells as previously described. RESULTS: Chronic exposure of cells to 200 mM ethanol for 24 and 48 hr resulted in the gradual re-distribution of RACK1 out of the nucleus. It is interesting to note that acute ethanol re-challenge immediately after chronic treatment did not result in RACK1 translocation to the nucleus, and nuclear compartmentalization of RACK1 in response to acute ethanol was detected only after 24 hr of withdrawal. Similar patterns were obtained for c-fos expression. Chronic exposure to ethanol did not result in an increase in mRNA or protein levels of c-fos. Furthermore, acute ethanol exposure did not increase c-fos protein levels in cells that were first treated chronically with ethanol. However, transduction of exogenous RACK1 expressed as a Tat-fusion protein was able to rescue c-fos mRNA expression after chronic ethanol exposure. CONCLUSIONS: Our data suggest that RACK1 nuclear compartmentalization and ethanol-induced c-fos expression are transient and are desensitized to ethanol during prolonged exposure to high concentrations. The desensitization is temporary, and RACK1 can respond to acute ethanol treatment after a 24-hr withdrawal period. Our data further suggest that the altered compartmentalization of RACK1 leads to differences in c-fos expression upon acute or chronic exposure to ethanol. In summary, RACK1 is an important molecular mediator of the acute and chronic actions of ethanol on the expression of c-fos. These findings could have implications for the molecular signaling pathways leading to pathologic states associated with alcoholism, including toxicity.

Adaptation, Physiological↗

PubMed2003

Fyn kinase and NR2B-containing NMDA receptors regulate acute ethanol sensitivity but not ethanol intake or conditioned reward.

BACKGROUND: The tyrosine kinase Fyn previously has been shown to play a key role in mediating acute tolerance to ethanol. Recently, we found that the compartmentalization of Fyn to the NR2B subunit of the NMDA receptor (NMDAR) in the hippocampus regulates Fyn phosphorylation of NR2B in response to ethanol, which mediates the acute tolerance of NMDAR to ethanol inhibition in hippocampal slices. In this study we determined, first, whether acute tolerance to ethanol inhibition is mediated via NR2B-containing NMDARs in vivo and, second, whether the increase in acute sensitivity to ethanol in the Fyn-/- mice influences ethanol consumption or ethanol's conditioned rewarding effects. METHODS: A loss of righting reflex test was used to study the acute/sedative effects of ethanol after intraperitoneal injections of sedative doses of ethanol. Conditioned place preference was used to study the rewarding properties of ethanol. The two-bottle choice protocol was used to measure oral ethanol self-administration and preference as described previously. RESULTS: We found that systemic injection of the NR2B-containing NMDAR selective antagonist, ifenprodil, abolished the differences between Fyn+/+ and Fyn-/- mice in sensitivity to the acute sedative effects of ethanol. Moreover, we found that Fyn-/- and Fyn+/+ mice did not differ in their voluntary ethanol consumption or in the rewarding properties of ethanol. CONCLUSIONS: Our results suggest that the interaction between Fyn and NR2B mediates the acute sedative effects of ethanol, and that alteration in acute ethanol sensitivity does not necessarily correlate with levels of ethanol consumption or the rewarding properties of ethanol.

Alcohol Drinking↗

PubMed2002

NMDA receptor function is regulated by the inhibitory scaffolding protein, RACK1.

Phosphorylation regulates the function of ligand-gated ion channels such as the N-methyl d-aspartate (NMDA) receptor. Here we report a mechanism for modulation of the phosphorylation state and function of the NMDA receptor via an inhibitory scaffolding protein, RACK1. We found that RACK1 binds both the NR2B subunit of the NMDA receptor and the nonreceptor protein tyrosine kinase, Fyn. RACK1 inhibits Fyn phosphorylation of NR2B and decreases NMDA receptor-mediated currents in CA1 hippocampal slices. Peptides that disrupt the interactions between RACK1, NR2B, and Fyn induce phosphorylation and potentiate NMDA receptor-mediated currents. Therefore, RACK1 is a regulator of NMDA receptor function and may play a role in synaptic plasticity, addiction, learning, and memory.

Amino Acid Sequence↗

PubMed2002

Ethanol induces gene expression via nuclear compartmentalization of receptor for activated C kinase 1.

Scaffolding proteins such as receptor for activated C kinase (RACK) 1 are involved in the targeting of signaling proteins and play an important role in the regulation of signal transduction cascades. Recently, we found that in cultured cells and in vivo, acute ethanol exposure induces the nuclear compartmentalization of RACK1. To elucidate a physiological role for nuclear RACK1, the Tat protein transduction system was used to transduce RACK1 and RACK1-derived fragments into C6 glioma cells. We found that nuclear RACK1 is mediating the induction of the immediate early gene c-fos expression induced by ethanol. First, transduction of full-length RACK1 (Tat-RACK1) resulted in the induction of c-fos expression and enhancement of ethanol activities. Second, we determined that the C terminus of RACK1 (Tat-RACK1DeltaN) is mediating transcription. Third, we identified a dominant negative fragment of RACK1 that inhibited the nuclear compartmentalization of endogenous RACK1 and inhibited ethanol-induction of c-fos mRNA and protein expression. Last, acute exposure to ethanol or transduction of full-length Tat-RACK1 resulted in an increase in mRNA levels of an activator protein 1 site-containing gene, PAC1 (pituitary adenylate cyclase-activating polypeptide receptor type I), suggesting that nuclear RACK1 is involved in the regulation of the expression of genes that are altered upon acute ethanol treatment. These results may therefore have important implications for the study of alcohol addiction.

Animals↗

PubMedDate not supplied

Visual, auditory and bimodal activity in the banks of the lateral suprasylvian sulcus in the cat.

In addition to visually driven cells we found within the lateral suprasylvian visual cortex of cats a considerable number of auditory and/or bimodal cells. Most of the visually driven cells were direction and orientation selective with responses that were neither highly stimulus time locked nor very stable. Most of the auditory responses were also not very stable, had relatively high thresholds, and were readily habituated. Previous studies have suggested that populations of cells within the lateral suprasylvian area are specialized for the analysis of optic flow fields (Rauscheker et al., 1987; Sherk et al., 1995). Given that a remarkable proportion of cells within this area can be also driven by auditory stimuli, we hypothesize that the 'optic flow' model may be extended to the bimodal domain rather than restricted to visual clues only. This, however, remains to be corroborated experimentally.

Acoustic Stimulation↗