Allostatic Changes in the cAMP System Drive Opioid-Induced Adaptation in Striatal Dopamine Signaling

Abstract

Opioids are powerful addictive agents that alter dopaminergic influence on reward signaling in medium spiny neurons (MSNs) of the nucleus accumbens. Repeated opioid exposure triggers adaptive changes, shifting reward valuation to the allostatic state underlying tolerance. However, the cellular substrates and molecular logic underlying such allostatic changes are not well understood. Here, we report that the plasticity of dopamine-induced cyclic AMP (cAMP) signaling in MSNs serves as a cellular substrate for drug-induced allostatic adjustments. By recording cAMP responses to optically evoked dopamine in brain slices from mice subjected to various opioid exposure paradigms, we define profound neuronal-type-specific adaptations. We find that opioid exposure pivots the initial hyper-responsiveness of D1-MSNs toward D2-MSN dominance as dependence escalates. Presynaptic dopamine transporters and postsynaptic phosphodiesterases critically enable cell-specific adjustments of cAMP that control the balance between opponent D1-MSN and D2-MSN channels. We propose a quantitative model of opioid-induced allostatic adjustments in cAMP signal strength that balances circuit activity.

Keywords: GPCR; addiction; cAMP; dopamine; nucleus accumbens; opioid; plasticity; striatum.

Figure 1.. A Single Acute Exposure to Morphine Induces Changes in cAMP Signaling in the NAc

(A) Experimental approach utilizing CAMPER brain slices. (B) cAMP responses recorded following optical stimulation. (C) Max amplitude cAMP change. Naive D1-MSN 90.3 ± 5.5 nM cAMP (n = 51 neurons; 7 mice) versus acute D1-MSN 68.7 ± 5.2 nM cAMP (n = 41 neurons; 6 mice); t test p = 0.0066, Kolmogorov-Smirnov (KS) D = 0.3544. Naive D2-MSN 68.5 ± 6.1 nM cAMP (n = 47 neurons; 6 mice) versus acute D2-MSN 32.5 ± 2.9 nM cAMP (n = 40 neurons; 5 mice); t test p < 0.0001, KS D = 0.516. (D) Peak width quantification. Naive D1-MSN 6.3 ± 0.39 min (n = 51 neurons; 7 mice) versus acute D1-MSN 4.2 ± 0.25 min (n = 41 neurons; 6 mice); t test p = 0.0003, KS D = 0.4414. Naive D2-MSN 3.7 ± 0.27 min (n = 47 neurons; 6 mice) versus acute D2-MSN 2.5 ± 0.08 min (n = 40 neurons; 5 mice); t test p < 0.0001, KS D = 0.5883. (E) cAMP responses from 20-Hz stimulation at 3′ inter-stimulation intervals (ISIs). (F) cAMP responses from 20-Hz stimulation at 1′ ISIs.

Figure 2.. Morphine Exposure Induces Allostatic Changes in cAMP Signaling

(A) Morphine administration paradigms. (B) cAMP response from optical stimulation. (C) Max amplitude cAMP change. Acute D1-MSN 68.7 ± 5.2 nM cAMP (n = 41 neurons; 6 mice) versus tolerance D1-MSN 58 ± 4.5 nM cAMP (n = 39 neurons; 4 mice); t test p = 0.275, KS D = 0.2226. Acute D2-MSN 32.5 ± 2.9 nM cAMP (n = 40 neurons; 5 mice) versus tolerance D2-MSN 80 ± 5.4 nM cAMP (n = 46 neurons; 5 mice); t test p < 0.0001, KS D = 0.7261. (D) Response duration. Acute D1-MSN 4.2 ± 0.25 min (n = 41 neurons; 6 mice) versus tolerance D1-MSN 5.6 ± 0.19 min (n = 39 neurons; 4 mice); t test p < 0.0001, KS D = 0.6048. Acute D2-MSN 2.5 ± 0.08 min (n = 40 neurons; 5 mice) versus tolerance D2-MSN 2.4 ± 0.07 min (n = 46 neurons; 5 mice); t test p = 0.5208, KS D = 0.1761. (E) cAMP response from optical stimulation at 3′ or 1′ ISIs. (F) cAMP response from optical stimulation. (G) Max amplitude cAMP change. Naive D1-MSN 90.3 ± 5.5 nM cAMP (n = 51 neurons; 7 mice) versus abstinence D1-MSN 29.1 ± 4.9 nM cAMP (n = 35 neurons; 6 mice); t test p < 0.0001, KS D = 0.702. Naive D2-MSN 68.5 ± 6.1 nM cAMP (n = 47 neurons; 6 mice) versus abstinence D2-MSN 84.5 ± 7.2 nM cAMP (n = 38 neurons; 5 mice); t test p = 0.3572, KS D = 0.2021. (H) Response duration. Naive D1-MSN 6.3 ± 0.39 min (n = 51 neurons; 7 mice) versus abstinence D1-MSN 4.1 ± 0.29 min (n = 35 neurons; 6 mice); t test p = 0.0009, KS D = 0.4314. Naive D2-MSN 3.7 ± 0.27 min (n = 47 neurons; 6 mice) versus abstinence D2-MSN 2.7 ± 0.30 min (n = 38 neurons; 5 mice); t test p = 0.0641, KS D = 0.2861. (I) cAMP response from optical stimulation at 3′ or 1′ ISIs. Gray/purple line indicates initial response peak. Red line indicates max response.

Figure 3.. Dopamine Transporter and Phosphodiesterases Regulate Magnitude and Duration of cAMP Signaling

(A) DAT inhibition schematic. (B) cAMP response from optical stimulation. (C) Max amplitude cAMP change. D1-MSN ACSF 90.3 ± 5.5 nM cAMP (n = 51 neurons; 7 mice) versus D1-MSN DAT block 139.9 ± 4.8 nM cAMP (n = 38 neurons; 4 mice); t test p < 0.0001, KS D = 0.4892. D2-MSN ACSF 68.5 ± 6.1 nM cAMP (n = 47 neurons; 6 mice) versus D2-MSN DAT block 45.0 ± 4.2 nM cAMP (n = 49 neurons; 4 mice); t test p = 0.0030, KS D = 0.3678. (D) Response duration. D1-MSN ACSF 3.0 ± 0.29 min (n = 51 neurons; 7 mice) versus D1-MSN DAT block 4.5 ± 0.49 min (n = 38 neurons; 4 mice); t test p = 0.0052, KS D = 0.3700. D2-MSN ACSF 2.0 ± 0.25 min (n = 47 neurons; 6 mice) versus D2-MSN DAT block 3.5 ± 0.28 min (n = 49 neurons; 4 mice); t test p = 0.0010, KS D = 0.3969. (E) PDE inhibition schematic. (F) cAMP response from optical stimulation. (G) Max amplitude cAMP change. D1-MSN ACSF 90.3 ± 5.5 nM cAMP (n = 51 neurons; 7 mice) versus D1-MSN PDE block 104.8 ± 7.2 nM cAMP (n = 33 neurons; 4 mice); t test p = 0.2101, KS D = 0.2371. D2-MSN ACSF 68.5 ± 6.1 nM cAMP (n = 47 neurons; 6 mice) versus D2-MSN PDE block 28.3 ± 3.1 nM cAMP (n = 44 neurons; 4 mice); t test p < 0.0001, KS D = 0.5629. (H) Response duration. D1-MSN ACSF 3.0 ± 0.29 min (n = 51 neuron; 7 mice) versus D1-MSN PDE block 14.5 ± 1.10 min (n = 33 neurons; 4 mice); t test p < 0.0001, KS D = 0.8717. D2-MSN ACSF 2.0 ± 0.25 min (n = 47 neurons; 6 mice) versus D2-MSN PDE block 3.2 ± 0.34 min (n = 44 neurons; 4 mice); t test p = 0.0307, KS D = 0.3032.

Figure 4.. Acute Morphine Modulates DAT and PDE Regulation of cAMP Signaling

(A) DAT inhibition schematic. (B) cAMP response from optical stimulation. (C) Max amplitude cAMP change; gray line indicates mean Naive (Saline) DAT Block. D1-MSN ACSF 68.7 ± 5.2 nM cAMP (n = 41 neurons; 6 mice) versus D1-MSN DAT Block 107.4 ± 5.8 nM cAMP (n = 36 neurons; 4 mice); t test p < 0.0001, KS D = 0.5549. D2-MSN ACSF 32.5 ± 2.9 nM cAMP (n = 40 neurons; 5 mice) versus D2-MSN DAT Block 26.6 ± 3.3 nM cAMP (n = 42 neurons; 4 mice); t test p = 0.0766, KS D = 0.2821. (D) Response duration; gray line indicates mean naive (saline) DAT block. D1-MSN ACSF 2.9 ± 0.40 min (n = 41 neurons; 6 mice) versus D1-MSN DAT block 5.1 ± 0.44 min (n = 36 neurons; 4 mice); t test p = 0.0014, KS D = 0.4356. D2-MSN ACSF 1.9 ± 0.32 min (n = 40 neurons; 5 mice) versus D2-MSN DAT block 2.9 ± 0.36 min (n = 42 neurons; 4 mice); t test p = 0.0383, KS D = 0.3107. (E) PDE inhibition schematic. (F) cAMP response from optical stimulation. (G) Max amplitude cAMP change; gray line indicates mean naive (saline) PDE block. D1-MSN ACSF 68.7 ± 5.2 nM cAMP (n = 41 neurons; 6 mice) versus D1-MSN PDE block 113.8 ± 4.8 nM cAMP (n = 38 neurons; 4 mice); t test p < 0.0001, KS D = 0.6733. D2-MSN ACSF 32.5 ± 2.9 nM cAMP (n = 40 neurons; 5 mice) versus D2-MSN PDE block 4.9 ± 0.39 nM cAMP (n = 45 neurons; 4 mice); t test p < 0.0001, KS D = 0.8778. (H) Response duration; gray line indicates mean naive (saline) PDE block. D1-MSN ACSF 2.9 ± 0.40 min (n = 41 neurons; 6 mice) versus D1-MSN PDE block 12.3 ± 1.08 min (n = 38 neurons; 4 mice); t test p < 0.0001, KS D = 0.7015. D2-MSN ACSF 1.9 ± 0.32 min (n = 40 neurons; 5 mice) versus D2-MSN PDE block width, not detected (n = 45 neurons; 4 mice).

Figure 5.. Acute Morphine Modulates Frequency Tuning of Dopaminergic Responses

(A) Experimental schematic. (B) cAMP responses varying frequencies of optical stimulation. (C) Max amplitude cAMP change. (D) Stimulation frequency that generated half of the max amplitude change. D1-MSN saline 8.76 ± 1.09 Hz versus D1-MSN morphine 12.56 ± 1.22 Hz; t test p = 0.0421. D2-MSN saline 13.22 ± 1.12 Hz versus D2-MSN morphine 11.16 ± 0.88 Hz; t test p = 0.1628.

Figure 6.. Opioid-Induced Plasticity Modulates Frequency Tuning through the DAT and PDEs

(A) D1-MSN response to optical stimulation during DAT inhibition. (B) Max amplitude cAMP change in D1-MSN. (C) Stimulation frequency that generated half of the max amplitude change. D1-MSN naive ACSF 8.76 ± 1.09 Hz, D1-MSN naive DAT block 4.97 ± 0.49, D1-MSN opioid ACSF 12.56 ± 1.22 Hz, D1-MSN opioid DAT block 12.53 ± 1.09 Hz. (D) D2-MSN response to optical stimulation during DAT inhibition. (E) Max amplitude cAMP change in D2-MSN. (F) Stimulation frequency that generated half of the max amplitude change. D2-MSN naive ACSF 13.22 ± 1.12 Hz, D2-MSN naive DAT block 10.45 ± 0.87, D2-MSN opioid ACSF 11.16 ± 0.88 Hz, D2-MSN opioid DAT block 6.05 ± 0.98 Hz. (G) D1-MSN response to optical stimulation during PDE inhibition. (H) Max amplitude cAMP change in D1-MSN. (I) Stimulation frequency that generated half of the max amplitude change. D1-MSN naive ACSF 8.76 ± 1.09 Hz, D1-MSN naive PDE block 3.43 ± 0.42, D1-MSN opioid ACSF 12.56 ± 1.22 Hz, D1-MSN opioid PDE block 11.71 ± 0.88 Hz. (J) D2-MSN response to optical stimulation during PDE inhibition. (K) Max amplitude cAMP change in D2-MSN. (L) Stimulation frequency that generated half of the max amplitude change. D2-MSN naive ACSF 13.22 ± 1.12 Hz, D2-MSN naive PDE block 9.84 ± 0.92, D2-MSN opioid ACSF 11.16 ± 0.88 Hz, D2-MSN opioid PDE block, not detected. Two-way ANOVA; ns p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Figure 7.. cAMP Signaling Activity between MSN Channels Calculates Shifts in Allostatic States

(A) Principle component analysis on correlations utilizing basal cAMP, signal duration, cAMP response amplitude, and cAMP rebound amplitude during phases of opioid exposure. Vectors represent mean population directionality from zero. (B) Aggregate Z score of signaling parameters (basal cAMP, response amplitude, and signal duration) from each phase of opioid exposure normalized to naive condition. (C) Z score ratio of D1:D2 from each phase of opioid exposure.