AbstractThis study examines the effect of intravenous self‐administration (SA) of either heroin or the cannabinoid receptor agonist WIN 55,212‐2 on levels and functionality of µ‐opioid (MOR) and CB1‐cannabinoid receptors (CB1R) in reward‐related brain areas, such as the prefrontal cortex (PFC), nucleus accumbens (NAc), caudate putamen (CP), hippocampus (Hippo), amygdala (Amy), hypothalamus (Hypo) and ventral tegmental area (VTA). [3H]DAMGO and [3H]CP‐55,940 autoradiography and agonist‐stimulated [35S]GTPγS binding were performed on brain sections of rats firmly self‐administering heroin or WIN 55,212‐2. Animals failing to acquire heroin or cannabinoid SA behaviour as well as drug‐naïve animals never exposed to experimental apparatus or procedure (home‐control group) were used as controls. With respect to control groups, which displayed very similar values, rats SA heroin showed increased MOR binding in the NAc (+174%), CP (+165%), Hippo (+121%), VTA (+175%), an enhanced CB1R density localized in the Amy (+147%) and VTA (+37%), and a widespread increased CB1 receptor functionality in the PFC (+95%), NAc (+313%), CP (+265%), Hippo (+38%), Amy (+221%). In turn, cannabinoid SA differently modulates CB1R binding in the Amy (+47%), Hypo (+94%), Hippo (−23%), VTA (−15%), and increases MOR levels (PFC: +124%; NAc: +68%; CP: +80%; Hippo: +73%; Amy: +99%) and efficiency (Hippo: +518%; Amy: +173%; Hypo: +188%). These findings suggest that voluntary chronic intake of opioids or cannabinoids induces reciprocal but differential regulation of MORs and CB1Rs density and activity in brain structures underlying drug‐taking and drug‐seeking behaviour, which could represent long‐term neuroadaptations contributing to the development of drug addiction and dependence.