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Submitted to: Research in Veterinary Science
Publication Type: Peer Reviewed Journal Publication Acceptance Date: 4/30/2017 Publication Date: 5/8/2017 Citation: Green, B.T., Lee, S.T., Welch, K.D., Cook, D. 2017. Anagyrine desensitization of peripheral nicotinic acetylcholine receptors. A potential biomarker of quinolizidine alkaloid teratogenesis in cattle.. Research in Veterinary Science. 115:195-200. doi: 10.1016/j.rvsc.2017.04.019. Interpretive Summary: Certain lupine species in the Western United States cause crooked calf syndrome. One alkaloid in particular, Anagyrine is particularly problematic in cattle. This alkaloid which is a quinolizidine alkaloid has been proposed to be metabolized to a piperidine alkaloid that inhibits fetal movement, the putative mechanism behind crooked calf syndrome. In this research using two cell lines we tested an alternative hypothesis that anagyrine directly desensitizes fetal muscle-type nAChR. Unexpectedly, anagyrine more potently desensitized autonomic nAChR suggesting a novel mechanism of teratogenesis by anagyrine. Moreover, serum anagyrine concentrations may be a potential biomarker for lupine teratogenicity in cattle. Technical Abstract: Anagyrine, a teratogenic quinolizidine alkaloid found in certain Lupinus spp., has been proposed to undergo metabolism by pregnant cattle to a piperidine alkaloid which acts inhibit fetal movement, the putative mechanism behind crooked calf syndrome. The objective of this study was to test the hypothesis that the quinolizidine alkaloids anagyrine but not lupanine or sparteine can directly, without metabolism, desensitize nicotinic acetylcholine receptors (nAChR) in a cell culture model. SH-SY5Y cells expressing autonomic nAChR, and TE-671 cells expressing fetal muscle-type nAChR were exposed to quinolizidines or dimethyphenylpiperazinium (DMPP) in log10 molar increments from 10 nM to 100 µM and then to a fixed concentration of acetylcholine (ACh) (10 µM for SH-SY5Y cells and 1 µM for TE-671 cells) and the responses measured with a membrane potential sensing dye to assess nAChR desensitization. The selective ganglionic nAChR agonist DMPP used as a positive control, was a potent activator and desensitizer of nAChR expressed by SH-SY5Y cells. Lupanine was a weak agonist and desensitizer in SH-SY5Y cells and sparteine was without effect. Anagyrine acted as a partial agonist in both cell lines with EC50 values of 4.2 and 231 µM in SH-SY5Y and TE-671 cells, respectively. Anagyrine was a desensitizer of nAChR with DC50 values of 6.9 and 139 µM in SH-SY5Y and TE-671 cells, respectively. These results confirm the hypothesis that anagyrine is a potent and effective desensitizer of nAChR, and that anagyrine can directly, without metabolism, desensitize nAChR. Moreover, serum anagyrine concentrations may be a potential biomarker for lupine teratogenicity in cattle. |
