Pituitary adenylate cyclase-activating polypeptide (PACAP) 38 and PACAP4-6 are neuroprotective through inhibition of NADPH oxidase: potent regulators of microglia-mediated oxidative stress.
Microglial activation is implicated in the progressive nature of numerous neurodegenerative diseases, including Parkinson's disease. Using primary rat mesencephalic neuron-glia cultures, we found that pituitary adenylate cyclase-activating polypeptide (PACAP) 38, PACAP27, and its internal peptide, Gly-Ile-Phe (GIF; PACAP4-6), are neuroprotective at 10(-13) M against lipopolysaccharide (LPS)-induced dopaminergic (DA) neurotoxicity, as determined by [(3)H]DA uptake and the number of tyrosine hydroxylase-immunoreactive neurons. PACAP38 and GIF also protected against 1-methyl-4-phenylpyridinium(+)-induced neurotoxicity but only in cultures containing microglia. PACAP38 and GIF ameliorated the production of microglia-derived reactive oxygen species (ROS), where both LPS- and phorbol 12-myristate 13-acetate-induced superoxide and intracellular ROS were inhibited. The critical role of NADPH oxidase for GIF and PACAP38 neuroprotection against LPS-induced DA neurotoxicity was demonstrated using neuron-glia cultures from mice deficient in NADPH oxidase (PHOX(-/-)), where PACAP38 and GIF reduced tumor necrosis factor alpha production and were neuroprotective only in PHOX(+/+) cultures and not in PHOX(-/-) cultures. Pretreatment with PACAP6-38 (3 microM; PACAP-specific receptor antagonist) was unable to attenuate PACAP38, PACAP27, or GIF (10(-13) M) neuroprotection. PACAP38 and GIF (10(-13) M) failed to induce cAMP in neuronglia cultures, supporting that the neuroprotective effect was independent of traditional high-affinity PACAP receptors. Pharmacophore analysis revealed that GIF shares common chemical properties (hydrogen bond acceptor, positive ionizable, and hydrophobic regions) with other subpicomolar-acting compounds known to inhibit NADPH oxidase: naloxone, dextromethorphan, and Gly-Gly-Phe. These results indicate a common high-affinity site of action across numerous diverse peptides and compounds, revealing a basic neuropeptide regulatory mechanism that inhibits microglia-derived oxidative stress and promotes neuron survival.
Yang
Sufen
S
Laboratory of Pharmacology and Chemistry, National Institute of Environmental Health Sciences, National Institutes of Health, MD F1-01, P.O. Box 12233, Research Triangle Park, NC 27709, USA.
Yang
Jun
J
Yang
Zhengqin
Z
Chen
Posee
P
Fraser
Alison
A
Zhang
Wei
W
Pang
Hao
H
Gao
Xi
X
Wilson
Belinda
B
Hong
Jau-Shyong
JS
Block
Michelle L
ML
D000818:Animals; D000242:Cyclic AMP / Q000096:biosynthesis; D005260:Female; D008070:Lipopolysaccharides / Q000633:toxicity; D017628:Microglia / Q000187:drug effects* / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors* / Q000502:physiology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009842:Oligopeptides / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide / Q000494:pharmacology*; D051381:Rats; D011916:Rats, Inbred F344
D008070:Lipopolysaccharides; D018696:Neuroprotective Agents; D009842:Oligopeptides; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D000242:Cyclic AMP; D019255:NADPH Oxidases