Intramural Research Program, National Institute on Drug Abuse-National Institutes of Health, Baltimore, Maryland, 21224
Intramural Research Program, National Institute on Drug Abuse-National Institutes of Health, Baltimore, Maryland, 21224.
J Neurosci. 2023 Mar 8;43(10):1692-1713. doi: 10.1523/JNEUROSCI.2049-22.2023. Epub 2023 Jan 30.
The brain µ-opioid receptor (MOR) is critical for the analgesic, rewarding, and addictive effects of opioid drugs. However, in rat models of opioid-related behaviors, the circuit mechanisms of MOR-expressing cells are less known because of a lack of genetic tools to selectively manipulate them. We introduce a CRISPR-based knock-in transgenic rat that provides cell type-specific genetic access to MOR-expressing cells. After performing anatomic and behavioral validation experiments, we used the knock-in rats to study the involvement of NAc MOR-expressing cells in heroin self-administration in male and female rats. Using RNAscope, autoradiography, and FISH chain reaction (HCR-FISH), we found no differences in expression in NAc, dorsal striatum, and dorsal hippocampus, or MOR receptor density (except dorsal striatum) or function between knock-in rats and wildtype littermates. HCR-FISH assay showed that is highly coexpressed with (95%-98%). There were no genotype differences in pain responses, morphine analgesia and tolerance, heroin self-administration, and relapse-related behaviors. We used the Cre-dependent vector AAV1-EF1a-Flex-taCasp3-TEVP to lesion NAc MOR-expressing cells. We found that the lesions decreased acquisition of heroin self-administration in male rats and had a stronger inhibitory effect on the effort to self-administer heroin in female rats. The validation of an knock-in rat enables new strategies for understanding the role of MOR-expressing cells in rat models of opioid addiction, pain-related behaviors, and other opioid-mediated functions. Our initial mechanistic study indicates that lesioning NAc MOR-expressing cells had different effects on heroin self-administration in male and female rats. The brain µ-opioid receptor (MOR) is critical for the analgesic, rewarding, and addictive effects of opioid drugs. However, in rat models of opioid-related behaviors, the circuit mechanisms of MOR-expressing cells are less known because of a lack of genetic tools to selectively manipulate them. We introduce a CRISPR-based knock-in transgenic rat that provides cell type-specific genetic access to brain MOR-expressing cells. After performing anatomical and behavioral validation experiments, we used the knock-in rats to show that lesioning NAc MOR-expressing cells had different effects on heroin self-administration in males and females. The new rats can be used to study the role of brain MOR-expressing cells in animal models of opioid addiction, pain-related behaviors, and other opioid-mediated functions.
脑μ阿片受体（MOR）对于阿片类药物的镇痛、奖赏和成瘾作用至关重要。然而，在与阿片类药物相关的行为的大鼠模型中，由于缺乏选择性操纵它们的遗传工具，MOR 表达细胞的回路机制知之甚少。我们引入了一种基于 CRISPR 的基因敲入转基因大鼠，为脑 MOR 表达细胞提供了细胞类型特异性的遗传访问。在进行解剖学和行为验证实验后，我们使用基因敲入大鼠来研究 NAc MOR 表达细胞在雄性和雌性大鼠海洛因自我给药中的作用。使用 RNAscope、放射自显影和 FISH 链反应（HCR-FISH），我们发现 NAc、背侧纹状体和背侧海马中的表达或 MOR 受体密度（除了背侧纹状体）或功能在基因敲入大鼠和野生型同窝仔鼠之间没有差异。HCR-FISH 检测显示，与 MOR 高度共表达（95%-98%）。在疼痛反应、吗啡镇痛和耐受、海洛因自我给药以及与复发相关的行为方面，没有基因型差异。我们使用 Cre 依赖性载体 AAV1-EF1a-Flex-taCasp3-TEVP 来损伤 NAc MOR 表达细胞。我们发现，损伤降低了雄性 大鼠海洛因自我给药的获得，并且对雌性 大鼠自我给予海洛因的努力有更强的抑制作用。基因敲入大鼠的验证使我们能够理解 MOR 表达细胞在大鼠阿片类药物成瘾、疼痛相关行为和其他阿片类药物介导的功能模型中的作用的新策略。我们的初步机制研究表明，损伤 NAc MOR 表达细胞对雄性和雌性大鼠的海洛因自我给药有不同的影响。
