Palmitic acid mediates hypothalamic insulin resistance by altering PKC-θ subcellular localization in rodents
Stephen C. Benoit, … , William L. Holland, Deborah J. Clegg
Stephen C. Benoit, … , William L. Holland, Deborah J. Clegg
Published August 10, 2009
Citation Information: J Clin Invest. 2009;119(9):2577-2589. https://doi.org/10.1172/JCI36714.
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Research Article

Palmitic acid mediates hypothalamic insulin resistance by altering PKC-θ subcellular localization in rodents

Abstract

Insulin signaling can be modulated by several isoforms of PKC in peripheral tissues. Here, we assessed whether one specific isoform, PKC-θ, was expressed in critical CNS regions that regulate energy balance and whether it mediated the deleterious effects of diets high in fat, specifically palmitic acid, on hypothalamic insulin activity in rats and mice. Using a combination of in situ hybridization and immunohistochemistry, we found that PKC-θ was expressed in discrete neuronal populations of the arcuate nucleus, specifically the neuropeptide Y/agouti-related protein neurons and the dorsal medial nucleus in the hypothalamus. CNS exposure to palmitic acid via direct infusion or by oral gavage increased the localization of PKC-θ to cell membranes in the hypothalamus, which was associated with impaired hypothalamic insulin and leptin signaling. This finding was specific for palmitic acid, as the monounsaturated fatty acid, oleic acid, neither increased membrane localization of PKC-θ nor induced insulin resistance. Finally, arcuate-specific knockdown of PKC-θ attenuated diet-induced obesity and improved insulin signaling. These results suggest that many of the deleterious effects of high-fat diets, specifically those enriched with palmitic acid, are CNS mediated via PKC-θ activation, resulting in reduced insulin activity.

Authors

Stephen C. Benoit, Christopher J. Kemp, Carol F. Elias, William Abplanalp, James P. Herman, Stephanie Migrenne, Anne-Laure Lefevre, Céline Cruciani-Guglielmacci, Christophe Magnan, Fang Yu, Kevin Niswender, Boman G. Irani, William L. Holland, Deborah J. Clegg

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Figure 7

Arcuate-specific knockdown of PKC-θ attenuates diet-induced obesity.

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Arcuate-specific knockdown of PKC-θ attenuates diet-induced obesity. (A)...
(A) AAV PKC-θ shRNA knocked down expression of PKC-θ in neuronal culture (lanes 1 and 3). (B) Attenuated diet-induced obesity in mice (n = 8–10/group), following bilateral arcuate infusion of 0.5 μl AAV PKC-θ shRNA or scrambled oligo AAV administration (data are mean ± SEM; *P < 0.05 relative to scrambled oligo control for body weight change). (C) Improved response to an i.p. glucose tolerance test (mean ± SEM; n = 8–10/group) in mice on the HFS diet, following AAV PKC-θ shRNA versus scrambled oligo AAV administration (*P < 0.05 relative to scrambled oligo AAV injections). (D) Improved insulin signaling (enhanced p-AKT 30 minutes following a peripheral injection of 5 U/kg) in mice (n = 4–5/group) on the HFS diet, following AAV PKC-θ shRNA versus scrambled oligo AAV administration (data are mean ± SEM; *P < 0.05 relative to control). (E) GFP immunoreactivity in the arcuate nucleus demonstrates transfection and expression of viral proteins at the site of injection. Original magnification, ×10 (left panel); ×20 (right panel). (F) Western blot analysis demonstrates a significant reduction (*P < 0.05) in PKC-θ protein levels, 2.5 weeks after AAV PKC-θ shRNA administration, only in the arcuate after AAV PKC-θ shRNA versus scrambled oligo AAV administration (quantification [mean ± SEM] from 2 separate Western blots; n = 4–5/group), (G) with no changes in PKC-θ in the remaining hypothalamus (quantification [mean ± SEM] from 2 separate Western blots; n = 4–5/group). (H and I) PKC-δ, a PKC isoform with high sequence homology to PKC-θ was not affected by viral injections (quantification [mean ± SEM] from 2 separate Western blots; n = 4–5/group). (J) The AAV PKC-θ shRNA versus scrambled oligo AAV administration had no effect on muscle levels of PKC-θ (quantification [mean ± SEM] from 2 separate Western blots; n = 4–5/group).