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. 2003 Nov 25;100(24):14379-84.
doi: 10.1073/pnas.2336093100. Epub 2003 Nov 6.

Addicting drugs utilize a synergistic molecular mechanism in common requiring adenosine and Gi-beta gamma dimers

Lina Yao  1 , Peidong Fan, Zhan Jiang, William S Mailliard, Adrienne S Gordon, Ivan Diamond
Affiliations

Addicting drugs utilize a synergistic molecular mechanism in common requiring adenosine and Gi-beta gamma dimers

Lina Yao et al. Proc Natl Acad Sci U S A. .

Abstract

The mesolimbic dopamine system and cAMP-dependent/protein kinase A (PKA) pathways are strongly implicated in addictive behaviors. Here we determine the role of dopamine D2 receptors (D2) in PKA signaling responses to delta-opioid (DOR) and cannabinoid (CB1) receptors. We find in NG108-15/D2 cells and in cultured primary neurons that a brief exposure to saturating concentrations of DOR and CB1 agonists increases cAMP, promotes PKA C alpha translocation and increases cAMP-dependent gene expression. Activation of PKA signaling is mediated by Gi-beta gamma dimers. Importantly, subthreshold concentrations of DOR or CB1 agonists with D2 agonists, which are without effect when added separately, together activate cAMP/PKA signaling synergistically. There is also synergy between DOR or CB1 with ethanol, another addicting agent. In all instances, synergy requires adenosine activation of adenosine A2 receptors and is mediated by beta gamma dimers. Synergy by this molecular mechanism appears to confer hypersensitivity to opioids and cannabinoids while simultaneously increasing the sensitivity of D2 signaling when receptors are expressed on the same cells. This mechanism may account, in part, for drug-induced activation of medium spiny neurons in the nucleus accumbens.

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Figures

Fig. 1.
Fig. 1.
Activation of DOR or CB1 promotes PKA Cα translocation and CRE-Luc expression in NG cells. (A) Cα translocation detected by immunostaining. NG cells were incubated with or without 1 μM DADLE (DAD), 2 μM Met, 10 μM UK, or 10 μM Carb or forskolin (FSK) (1 μM) for 10 min. Where indicated, cells were preincubated either with the DOR antagonist Nal (10 μM) or the CB1 antagonist AM (10 μM) for 30 min. Data represent at least three experiments. Staining intensity is indicated by the color bar (red indicates highest concentration) (scale bar, 10 μm; magnification, ×400). (B)Cα translocation detected by Western blots of nuclear (N), membrane (M), and cytosolic (C) fractions from treated cells. (C) Time course of cAMP production. Cells were incubated with or without 1 μM DADLE, 2 μM Met, 10 μM UK, or 10 μM Carb for the indicated times. cAMP was measured by RIA (17). Data are the mean ± SEM of four experiments. *, P < 0.05 compared with time 0 (one-way analysis of variance and Dunnett's test). cAMP levels in the absence of drugs did not change during the experiment. (D) Rp and PTX inhibit PKA Cα translocation. Cells were pretreated with 20 μM Rp for 1.5 h or PTX (50 ng/ml) overnight before incubation with DADLE or Met as in A. (E) βγ dimers are required for PKA Cα translocation. Cells were transfected with the βγ inhibitors Ad5βARK1 or Ad5QEHA or Ad5 vector control and incubated with or without 1 μM DADLE or 2 μM Met. (F) Activation of DOR or CB1 induces CRE-Luc expression. Cells were transiently transfected with a CRE-Luc construct, preincubated with buffer or Nal, AM, Rp, or PTX as above and then treated for 10 min with or without DADLE, Met, or forskolin. Luc was assayed 5 h after drug treatment. Data are the mean ± SEM of at least three experiments. *, P < 0.01 compared with control (one-way analysis of variance and Dunnett's test).
Fig. 2.
Fig. 2.
Synergy for PKA Cα translocation and CRE-Luc expression between subthreshold concentrations of DADLE or Met with NPA or ethanol. (A) Subthreshold concentrations of NPA (0.5 nM), DADLE (0.01 nM), Met (0.02 nM), or ethanol (E) (25 mM) that did not induce translocation alone were tested for synergy during a 10-min incubation (Top). Cells were also preincubated in Rp or QEHA (Middle and Bottom). (B) Cells were preincubated with or without 10 μM BW before testing for translocation synergy with subthreshold concentrations of NPA, DADLE, Met, or ethanol as in A. (C) Cells were incubated as in A, in the presence or absence of 1 unit/ml adenosine deaminase (ADA). (D) Cells transiently transfected with CRE-Luc were treated for 10 min as in A, and Luc was measured 5 h later. Data are the mean ± SEM of at least three experiments. *, P < 0.01 compared with control (one-way analysis of variance and Dunnett's test). (E) Cells were preincubated with (black bars) or without (red bars) BW (10 μM) for 1 h and then further incubated in 1 μM DADLE or 2 μM Met. Data are the mean ± SEM of at least three experiments.
Fig. 4.
Fig. 4.
Schematic representation of postsynaptic DOR, CB1, and D2 activation-induced PKA Cα translocation and CRE-mediated gene expression via βγ dimers. A central role for βγ subunits released from Gi/o is proposed for DOR, CB1, and D2. This diagram indicates synergy between subthreshold levels of DOR or CB1 (blue arrows) with D2 (red arrows). Synergy for PKA translocation and CRE-mediated gene expression is mediated by βγ dimers from Gi/o. Adenosine A2 activation via Gαs is required for synergy. We propose that colocalization of A2, D2, DOR, and CB1 on the same neurons, like in NAc, confers hypersensitivity to exogenous opioids, cannabinoids, and ethanol because of synergy. In addition, synergy promotes simultaneous hypersensitivity of postsynaptic D2 signaling that is characteristic of addicting drugs.
Fig. 3.
Fig. 3.
DOR or CB1 agonists induce PKA Cα translocation and CRE-Luc expression in rat PHN. (A) PHN were preincubated in the presence or absence of buffer, Nal, or AM, and then incubated in the presence or absence of 1 μM DADLE or 2 μM Met for 10 min. Cα is indicated by green staining, and the neuron-specific marker, NeuN, is indicated by red staining. (B) PHN were preincubated with or without Rp for 1.5 h, or overnight with or without PTX or QEHA before incubation with or without DADLE or Met as in A. (C) PHN were treated with subthreshold concentrations of DADLE, Met, NPA, or ethanol alone or in combination as in Fig. 2 A. Where indicated, the cells were preincubated for 1 h with 1,3-dipropyl-8 cyclopentylxanthine (DPCPX) (100 nM) or 3,7-dimethyl-1-(2-propynyl)xanthine (DMPX) (10 μM). Incubation with subthreshold concentrations of each agent alone was without effect.
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References

    1. Robbins, T. W. & Everitt, B. J. (1999) Nature 398, 567-570. - PubMed
    1. Nestler, E. J. (2001) Nat. Rev. Neurosci. 2, 119-128. - PubMed
    1. Hyman, S. E. & Malenka, R. C. (2001) Nat. Rev. Neurosci. 2, 695-703. - PubMed
    1. Albert, P. R. & Robillard, L. (2002) Cell. Signal. 14, 407-418. - PubMed
    1. Tang, W. J. & Gilman, A. G. (1991) Science 254, 1500-1503. - PubMed

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