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Injection of Toll-like receptor 4 siRNA into the ventrolateral periaqueductal gray attenuates withdrawal syndrome in morphine-dependent rats

Q F Liu, L Li, Y Q Guo, X Li, Z D Mou, X Wang, G Z Du


We assessed the role of the Toll-like receptor 4 (TLR4) gene in the ventrolateral periaqueductal gray (vlPAG) region of morphine-dependent rats on attenuating withdrawal syndrome, and regulating glutamic acid decarboxylase (GAD67), glutamic acid (Glu), and gamma-aminobutyric acid (GABA). After siRNA-mediated downregulation of TLR4, changes were observed in withdrawal behavior and downstream signaling molecules. Rats were injected into the vlPAG with TLR4 siRNA, followed by intraperitoneal injection of morphine for 5 consecutive days, and then naloxone, and the behavioral indices of morphine withdrawal were observed. ‘Wet-dog’ shakes, teeth chattering, and the total scores of withdrawal reactions were reduced. TLR4 expression and Glu levels were reduced, whereas GAD67 and GABA levels were increased. Overall, these findings indicate that modifying TLR4 gene expression in the vlPAG stimulates expression of the downstream signaling molecule, GAD67, which decreases Glu levels and increases GABA levels. This mechanism may explain the inhibition of withdrawal syndrome in morphine-dependent rats.


Toll-like receptor 4; morphine withdrawal syndrome; GAD67; GABA

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Alfonso-Loeches S., Pascual-Lucas M., Blanco A.M., Sanchez-Vera I., Guerri C. Pivotal role of TLR4 receptors in alcohol-induced neuroinflammation and brain damage. J. Neurosci., 30 (24): 8285-8295, 2010.

Bagley E. E. Opioid and GABAB receptors differentially couple to an adenylyl cyclase/protein kinase A downstream effector after chronic morphine treatment. Front. Pharmacol., 5:148, 2014.

Balosso S., Maroso M., Sanchez-Alavez M., Ravizza T., Frasca A., Bartfai T., Vezzani A. A novel non-transcriptional pathway mediates the proconvulsive effects of interleukin-1β. Brain, 131 (12): 3256-3265, 2008.

Bradford M. M. Determination of proteins from sediment samples. Pangaea, 1976.

Chen M., Zhao Y., Yang H., Luan W., Song J., Cui D., Dong Y., Lai B., Ma L., Zheng P. Morphine disinhibits glutamatergic input to VTA dopamine neurons and promotes dopamine neuron excitation. Elife, 4 e09275, 2015.

Eidson, L. N. and A. Z. Murphy. Blockade of Toll-like receptor 4 attenuates morphine tolerance and facilitates the pain relieving properties of morphine. J. Neurosci., 33 (40): 15952-15963, 2013.

Hao S.L., Hu J., Fink D. J. Transgene-mediated enkephalin expression attenuates signs of naloxone-precipitated morphine withdrawal in rats with neuropathic pain. Behav Brain Res., 30; 197(1): 84–89, 2009.

Hutchinson M.R., Northcutt A.L., Hiranita T., Wang X., Lewis S.S., Thomas J., van Steeg K., Kopajtic T.A., Loram L.C., Sfregola C., Galer E., Miles N.E., Bland S.T., Amat J., Rozeske R.R., Maslanik T., Chapman T.R., Strand K.A., Fleshner M., Bachtell R.K., Somogyi A.A., Yin H., Katz J.L., Rice K.C., Maier S.F., Watkins L.R. Opioid activation of toll-like receptor 4 contributes to drug reinforcement. J. Neurosci., 32 (33): 11187-11200, 2012.

Lan L.S., Ping Y.J., Na W.L., Miao J., Cheng Q.Q., Ni M.Z., Lei L., Fang L.C., Guang R.C., Jin Z., Wei L. Down-regulation of Toll-like receptor 4 gene expression by short interfering RNA attenuates bone cancer pain in a rat model. Mol. pain, 6 (1): 1, 2010.

Liu J., Yang A.R., Kelly T., Puche A., Esoga C., June H.L. Jr, Elnabawi A., Merchenthaler I., Sieghart W., June H.L Sr, Aurelian L. Binge alcohol drinking is associated with GABAA α2-regulated Toll-like receptor 4 (TLR4) expression in the central amygdala. Proc. Natl. Acad. Sci. U S A., 108 (11): 4465-4470, 2011.

Maroso M., Balosso S., Ravizza T., Liu J., Aronica E., Iyer A.M., Rossetti C., Molteni M., Casalgrandi M., Manfredi A.A., Bianchi M.E., Vezzani A. Toll-like receptor 4 and high-mobility group box-1 are involved in ictogenesis and can be targeted to reduce seizures. Nat. med., 16 (4): 413-419, 2010.

Wu F., Miao X.R., XU X.W., Sun Y.M. Intrathecal injection of siRNA targeting Toll-like receptor 4 reduces neuropathic pain in rat model of chronic constriction injury. Academic Journal of Second Military Medical University, 32 (4): 377-381, 2011.

Wu H.H. and Hou Y.N. Changes of Levels of Amino Acids in Brain Regions of Morphine Psychic

Dependent Rats. Pharmaceutical Journal of Chinese People's Liberation Army, 4 2005.

Xu Y and Xu M. Effects of r -aminobutyric acid on morphine withdrawal symptoms in rats and its possible mechanism. Chinese Journal of Drug Dependence, 15 (01): 21-23, 2006.

Yan X., Jiang E., Weng HR. Activation of toll like receptor 4 attenuates GABA synthesis and postsynaptic GABA receptor activities in the spinal dorsal horn via releasing interleukin-1 beta. J. neuroinflammation, 12 (1): 222, 2015.

Zamberletti E., Beggiato S., Steardo L. Jr., Prini P., Antonelli T., Ferraro L., Rubino T., Parolaro D. Alterations of prefrontal cortex GABAergic transmission in the complex psychotic-like phenotype induced by adolescent delta-9-tetrahydrocannabinol exposure in rats. Neurobio.l Dis., 63 (5): 35-47, 2014



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