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Science 338 (6103): 124-128

Copyright © 2012 by the American Association for the Advancement of Science

BDNF Is a Negative Modulator of Morphine Action

Ja Wook Koo1, Michelle S. Mazei-Robison1, Dipesh Chaudhury2, Barbara Juarez2, Quincey LaPlant1, Deveroux Ferguson1, Jian Feng1, Haosheng Sun1, Kimberly N. Scobie1, Diane Damez-Werno1, Marshall Crumiller1, Yoshinori N. Ohnishi3, Yoko H. Ohnishi4, Ezekiell Mouzon1, David M. Dietz5, Mary Kay Lobo6, Rachael L. Neve7, Scott J. Russo1, Ming-Hu Han1,2, and Eric J. Nestler1,2,*

1 Fishberg Department of Neuroscience and Friedman Brain Institute, Mount Sinai School of Medicine, New York, NY 10029, USA.
2 Department of Pharmacology and Systems Therapeutics, Mount Sinai School of Medicine, New York, NY 10029, USA.
3 Department of Pharmacology, Kurume University School of Medicine, Kurume, Fukuoka 830-0011, Japan.
4 Department of Medical Biophysics and Radiation Biology, Kyushu University, Higashi-ku, Fukuoka 812-8582, Japan.
5 Department of Pharmacology and Toxicology, State University of New York at Buffalo, Buffalo, NY 14214, USA.
6 Department of Anatomy and Neurobiology, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
7 Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.


Figure 1
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Fig. 1. Effects of BDNF-TrkB signaling within the VTA-NAc on morphine reward. Localized knockout (KO) of BDNF (A) or TrkB (B) from VTA neurons enhances morphine CPP [15 mg/kg, subcutaneous (sc)]. Student’s t test, *P < 0.05, n = 8 to 12 mice. (C) DAT-Cre/flTrkB (TrkBlx/lx;DATcre/wt) mice also displayed enhanced morphine CPP [10 mg/kg, intraperitoneal (ip)]. One-way analysis of variance (ANOVA), Fisher's protected least significant difference (PLSD) post-hoc test, *P < 0.05 compared to controls; #P < 0.05 compared with TrkBlx/lx;DATcre/wt mice, n = 6 to 11 mice. (D) A single infusion of BDNF into the VTA (0.25 μg per side) suppressed morphine CPP (15 mg/kg, sc). PBS, phosphate-buffered saline. t test, *P < 0.05, n = 7 or 8 mice. (E) Localized TrkB KO in the NAc and (F) intra-NAc BDNF infusion (1.0 μg per side) had no effect on morphine CPP (15 mg/kg, sc), n = 8 or 9 mice.

 

Figure 2
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Fig. 2. Regulation of VTA DA neuron excitability by morphine and BDNF. (A) Sample traces of in vivo recordings from VTA DA neurons from control (top), morphine-treated (middle), and BDNF+morphine–treated mice (bottom). (B to E) Morphine (25-mg pellet, sc; animals were analyzed 48 hours later) increases (B) basal firing rate, (C) burst firing rate, and (D) burst duration in VTA DA neurons, which were normalized by intra-VTA infusion of BDNF (0.25 μg per side). One-way ANOVA, Fisher's PLSD test, *P < 0.05, ***P < 0.001 compared with control; ##P < 0.01, ###P < 0.001 compared with the morphine group. n = 4 to 6 mice. (E) Sample traces of K+ conductance recorded from VTA DA neurons in brain slices from control, morphine-treated, and BDNF+morphine–treated mice. (F) Morphine treatment as in (A) significantly decreased both peak and sustained phases of K+ currents in VTA DA neurons, an effect that was reversed by BDNF. Two-way ANOVA, Fisher's PLSD test, *P < 0.05, **P < 0.01, ***P < 0.001 compared with control; #P < 0.05, ##P < 0.01, ###P < 0.001 compared with the morphine group. n = 5 to 9 mice. Localized BDNF KO from VTA increases (G) basal firing rate, (H) burst firing rate, and (I) burst duration in VTA DA neurons. t test, *P < 0.05, **P < 0.01 compared with AAV-GFP controls, n = 7 mice.

 

Figure 3
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Fig. 3. Modulation of morphine reward by optogenetic activation of DA terminals in the NAc. (A) Experimental model for optogenetic stimulation during morphine CPP. Mice were conditioned to a morphine/light chamber and a saline/no-light chamber for 30 min. 470-nm phasic light pulses (20 Hz, five pulses, 40 ms duration) were delivered during the 30-min conditioning session. (B to J) Immunostaining for ChR2-EYFP [(B), (E), and (H), green], DAT [(C), red], GAD67 [(F), red], and VGLUT2 [(I), red] in the NAc. (D), (G), and (J) Confocal microscopy shows that ChR2-EYFP puncta in NAc co-label for DAT, but not GAD67 or VGLUT2. Scale bar, 10 μm. (K) In vivo optogenetic stimulation of VTA DA nerve terminals in the NAc enhances morphine reward (10 mg/kg, ip) and prevents VTA BDNF-induced impairment of morphine reward, (L) whereas D1 receptor antagonism (SCH 23390, 1 μg) blocks light potentiation, and VTA BDNF-induced impairment, of morphine reward. t test, **P < 0.01, **P < 0.001, n = 7 to 11 mice.

 

Figure 4
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Fig. 4. Morphine-regulated NAc gene expression after VTA BDNF deletion: Identification of NAc mediators of BDNF-morphine interactions. Microarray analysis was performed on the NAc of sham- and morphine-pelleted mice under control or VTA BDNF knockdown conditions. (A) Heat map of up-regulated (red) or down-regulated (green) NAc genes upon knockdown of VTA BDNF. (B) Heat map of up- or down-regulated NAc genes by morphine regardless of knockdown of VTA BDNF. (C) Venn diagrams of genes that were regulated by morphine (red) or by knockdown of VTA BDNF (blue), and of genes that were regulated by morphine and knockdown of VTA BDNF in an interactive manner (green). (D to G) Alterations of sox11 [(D) and (E)] and gadd45g [(F) and (G)] expression in the NAc from a heat map of microarray analysis [(D) and (F)] and RT-PCR (RT-PCR) validation [(E) and (G)]. One-way ANOVA for RT-PCR validation, Fisher's PLSD test, {tau}P < 0.1, *P < 0.05, ***P < 0.001 compared to sham+AAV-GFP controls; $P < 0.1, #P < 0.05, ###P < 0.001 compared to sham+AAV-CreGFP mice, n = 9 to 12 mice. (H) Reduction of sox11 expression using AAV-shRNA-Sox11 increases morphine CPP (10 mg/kg, sc). Fisher's PLSD test, *P < 0.05 compared with AAV-GFP controls; #P < 0.05 compared with AAV-scrambled controls, n = 11 or 12 mice. (I) Sox11 overexpression in NAc using HSV-Sox11 decreases morphine CPP (15 mg/kg, sc). t test, *P < 0.05, n = 8 mice. (J) Enhancement of morphine reward (15 mg/kg, sc) induced by knockdown of VTA TrkB is counteracted by sox11 overexpression in the NAc. Fisher's PLSD test, *P < 0.05 compared with HSV-tomato (TMT) (NAc)+AAV-GFP (VTA) controls; #P < 0.05 compared with HSV-TMT (NAc)+AAV-CreGFP (VTA) mice. (K) Enhancement of morphine reward (12.5 mg/kg, sc) induced by knockdown of VTA TrkB is further enhanced by gadd45g overexpression in the NAc. Fisher's PLSD test, *P < 0.05 compared to HSV-GFP+AAV-GFP controls; #P < 0.05, ###P < 0.001 compared to HSV-Gadd45g+AAV-CreGFP.

 


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