Verapamil protects dopaminergic neuron damage through a novel anti-inflammatory mechanism by inhibition of microglial activation.
Verapamil has been shown to be neuroprotective in several acute neurotoxicity models due to blockade of calcium entry into neurons. However, the potential use of verapamil to treat chronic neurodegenerative diseases has not been reported. Using rat primary mesencephalic neuron/glia cultures, we report that verapamil significantly inhibited LPS-induced dopaminergic neurotoxicity in both pre- and post-treatment experiments. Reconstituted culture studies revealed that the presence of microglia was essential in verapamil-elicited neuroprotection. Mechanistic studies showed that decreased production of inflammatory mediators from LPS-stimulated microglia underlay neuroprotective property of verapamil. Further studies demonstrated that microglial NADPH oxidase (PHOX), the key superoxide-producing enzyme, but not calcium channel in neurons, is the site of action for the neuroprotective effect of verapamil. This conclusion was supported by the following two observations: 1) Verapamil failed to show protective effect on LPS-induced dopaminergic neurotoxicity in PHOX-deficient (deficient in the catalytic subunit of gp91(phox)) neuron/glia cultures; 2) Ligand binding studies showed that the binding of [(3)H]Verapamil onto gp91(phox) transfected COS7 cell membranes was higher than the non-transfected control. The calcium channel-independent neuroprotective property of verapamil was further supported by the finding that R(+)-verapamil, a less active form in blocking calcium channel, showed the same potency in neuroprotection, inhibition of pro-inflammatory factors production and binding capacity to gp91(phox) membranes as R(-)-verapamil, the active isomer of calcium channel blocker. In conclusion, our results demonstrate a new indication of verapamil-mediated neuroprotection through a calcium channel-independent pathway and provide a valuable avenue for the development of therapy for inflammation-related neurodegenerative diseases.
Neuropharmacology Section, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, USA.
Yuxin
Y
Liu
Yi-Ching
YC
Lo
Li
L
Qian
Fulton Tim
FT
Crews
Belinda
B
Wilson
Hui-Ling
HL
Chen
Hung-Ming
HM
Wu
Shih-Heng
SH
Chen
Ke
K
Wei
Ru-Band
RB
Lu
Syed
S
Ali
Jau-Shyong
JS
Hong
D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology* / Q000627:therapeutic use; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004298:Dopamine / Q000378:metabolism*; D005260:Female; D007249:Inflammation / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D017628:Microglia / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D018696:Neuroprotective Agents / Q000494:pharmacology* / Q000627:therapeutic use; D011247:Pregnancy; D051381:Rats; D011916:Rats, Inbred F344; D014700:Verapamil / Q000494:pharmacology*
D000893:Anti-Inflammatory Agents; D008070:Lipopolysaccharides; D018696:Neuroprotective Agents; D014700:Verapamil; D004298:Dopamine