Heparan sulfate: Resilience factor and therapeutic target for cocaine abuse

Abstract

Substance abuse is a pressing problem with few therapeutic options. The identification of addiction resilience factors is a potential strategy to identify new mechanisms that can be targeted therapeutically. Heparan sulfate (HS) is a linear sulfated polysaccharide that is a component of the cell surface and extracellular matrix. Heparan sulfate modulates the activity and distribution of a set of negatively charged signaling peptides and proteins - known as the HS interactome - by acting as a co-receptor or alternative receptor for growth factors and other signaling peptides and sequestering and localizing them, among other actions. Here, we show that stimulants like cocaine and methamphetamine greatly increase HS content and sulfation levels in the lateral hypothalamus and that HS contributes to the regulation of cocaine seeking and taking. The ability of the HS-binding neuropeptide glial-cell-line-derived neurotrophic factor (GDNF) to increase cocaine intake was potentiated by a deletion that abolished its HS binding. The delivery of heparanase, the endo-β-D-glucuronidase that degrades HS, accelerated the acquisition of cocaine self-administration and promoted persistent responding during extinction. Altogether, these results indicate that HS is a resilience factor for cocaine abuse and a novel therapeutic target for the treatment of cocaine addiction.

Figure 1

Stimulants increase heparan sulfate abundance and sulfation. (a) Cocaine and methamphetamine increase HS content. Total HS abundance was significantly increased by cocaine (Coc, n = 6) and methamphetamine (Meth, n = 5) in the LH, a brain region where the HS proteoglycan syndecan-3 was previously found to regulate cocaine intake. (b) Cocaine and methamphetamine significantly increase HS sulfation. *P < 0.05, ****P < 0.0001, significant difference from saline (Sal) control (n = 6; one-way ANOVA followed by Fisher’s LSD test). The data are expressed as mean + SEM.

Figure 2

Effect of lateral hypothalamus delivery of GDNF-AAV and ΔN2-GDNF-AAV on cocaine self-administration in wildtype and syndecan-3 knockout mice. A unit dose-response paradigm was used to probe the effects of GDNF heparin binding in syndecan-3 knockout and wildtype mice under an FR5 schedule of reinforcement. (a) GDNF-AAV or ΔN2-GDNF-AAV was injected in the LH in wildtype and syndecan-3 KO mice. (b) No significant effect of GDNF-AAV or ΔN2-GDNF viruses was observed in wildtype (WT) mice compared with the GFP control group (F _2,228 = 1.48, P > 0.05, two-way repeated-measures ANOVA). GFP, n = 12; GDNF, n = 15; ΔND2-GDNF, n = 14. (c) In syndecan-3 knockout mice, both the GDNF-AAV and ΔN2-GDNF-AAV viruses caused an upward shift of the cocaine unit dose-response curve compared with the GFP control group. The two-way repeated-measures ANOVA revealed a significant main effect of virus infusion (F _2,180 = 4.26, P = 0.024). GFP, n = 10; GDNF, n = 12; ΔND2-GDNF, n = 11. The post hoc test revealed a significant group difference between the ΔN2-GDNF-AAV group and GFP group at the doses of 0.075, 0.15, and 0.3 mg/kg/injection. (d) ΔN2-GDNF-AAV delivery increased cocaine intake compared with the GFP control group. The two-way repeated-measures ANOVA revealed a significant main effect of viral infusion (F _2,30 = 2.828, P = 0.075). GFP, n = 10; GDNF, n = 12; ΔND2-GDNF, n = 11. The post hoc test revealed a significant group difference between the ΔN2-GDNF-AAV group and GFP group at the doses of 0.6 and 1.2 mg/kg/injection. The data are expressed as mean ± SEM.

Figure 3

Delivery of recombinant AAV-heparanase in the LH facilitates the acquisition of cocaine self-administration in syndecan-3 knockout (KO) and wildtype (WT) mice and induces greater resistance to extinction in syndecan-3 KO mice. (a) Intra-LH heparanase-AAV virus facilitated the acquisition of cocaine self-administration in both wildtype (left) and syndecan-3 knockout (right) mice. The two-way repeated-measures ANOVA revealed significant main effects of repeated training (F _16,288 = 26.51, p < 0.0001) and heparanase-AAV administration (F _1,288 = 4.613, p = 0.046) and a significant interaction between these two factors (F _16,288 = 2.316, p = 0.003) in wildtype mice. Similarly, in knockout mice, there were significant main effects of repeated training (F _16,336 = 15.43, p < 0.0001) and heparanase-AAV administration (F _1,336 = 4.982, p = 0.037). The post hoc tests revealed significant differences on several training days between the corresponding groups (*P < 0.05, WT-GFP, n = 10; WT-Heparanase, n = 10; KO-GFP, n = 12; KO-Heparanase, n = 11). (b,c) The maintenance of cocaine self-administration was tested under a fixed-ratio 5 schedule of reinforcement (b), followed by a progressive-ratio schedule (c). Intra-LH heparanase-AAV did not significantly alter cocaine self-administration under either the FR or PR schedule. No significant main effects of heparanase-AAV administration were observed in either wildtype or knockout mice (P > 0.05). (d) The mice were then tested in daily extinction training. There was no significant main effect of heparanase-AAV in wildtype mice (F _1,450 = 0.398, p > 0.05). However, heparanase-AAV increased resistance to the extinction of cocaine seeking in syndecan-3 knockout mice, reflected by a significant main effect of heparanase-AAV treatment (F _1,450 = 5.33, p = 0.033; two-way repeated-measures ANOVA). The data are expressed as mean ± SEM.