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Role of somatostatin and muscarinic - acetylcholine system in 3NP-induced neurotoxicity in mutant Htt knock-in cultured striatal neuronal cells

University of British Columbia

Huntington’s disease (HD), is an inherited neurological disease with severe neuronal loss in the striatum. Previous studies have shown the selective sparing of somatostatin (SST) positive medium-sized aspiny interneurons and large cholinergic interneurons expressing choline acetyltransferase (ChAT) in the striatum. This suggests a crucial role of somatostatinergic and cholinergic neurotransmission in the pathophysiology of HD. The biological effects of SST in the central and peripheral systems are mediated by five different somatostatin receptors (SSTR1 - 5); whereas five metabotropic muscarinic receptors (M1R - M5R) mediate acetylcholine (ACh) functions. Whether SSTR and MR subtypes work in concert in HD is not known. In the present study, using STHdhQ7/7 (wt) cells and STHdhQ111/111 (mutant) cells and 3-Nitro propionic acid (3NP)-induced model of toxicity, first the expression levels of SSTR and MR subtypes were determined using the immunocytochemistry and Western blot analysis, second cell viability study and apoptosis were performed using MTT assay and Hoechst 33258 dye, third we determined the status of downstream signaling pathways including ERK1/2 and Akt upon treatment with SST and Carbachol (Carb) alone and in combination. STHdhQ7/7 and STHdhQ111/111 cells display moderate to strong expression and colocalization between M1R/M4R and SSTR2/4. Both STHdhQ7/7 and STHdhQ111/111 cells display significant changes in internalization and cell surface-expression of M1R/M4R and SSTR2/4 in a receptor- and treatment-specific manner. STHdhQ111/111 cells exhibit a slow proliferation rate than STHdhQ7/7 cells, and exhibit concentration- and time-dependent effect on the cell viability in the presence of 3NP. STHdhQ111/111 cells are more susceptible to 3NP-induced toxicity and apoptosis when compared to STHdhQ7/7 cells. In cell viability assay, agonist-treatment before and after 3NP-toxicity affords better protection than co-treatment. Consistent with receptor expression, cell proliferation and cell viability, time- and concentration-dependent regulation of signal transduction pathways- ERK1/2 and Akt attest the neuroprotective role of Carb and SST against 3NP-induced neurotoxicity. In conclusion, the results presented here shed a new light on the neuroprotective role of Carb and SST, and further enhance our understanding of functional interaction between MRs - SSTRs in an in vitro model of HD.

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2023-09-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/76243

Pharmaceutical Sciences

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/