Elevation of circulating fatty acids (FA) during fasting supports postprandial (PP) insulin secretion that is critical for glucose homeostasis and is impaired in diabetes. We tested our hypothesis that lipid droplet (LD) protein perilipin 5 (PLIN5) in β-cells aids PP insulin secretion by regulating intracellular lipid metabolism. We demonstrated that PLIN5 serves as an LD protein in human islets. In vivo, Plin5 and triglycerides were increased by fasting in mouse islets. MIN6 cells expressing PLIN5 (adenovirus [Ad]-PLIN5) and those expressing perilipin 2 (PLIN2) (Ad-PLIN2) had higher [(3)H]FA incorporation into triglycerides than Ad-GFP control, which support their roles as LD proteins. However, Ad-PLIN5 cells had higher lipolysis than Ad-PLIN2 cells, which increased further by 8-Br-cAMP, indicating that PLIN5 facilitates FA mobilization upon cAMP stimulation as seen postprandially. Ad-PLIN5 in islets enhanced the augmentation of glucose-stimulated insulin secretion by FA and 8-Br-cAMP in G-protein-coupled receptor 40 (GPR40)- and cAMP-activated protein kinase-dependent manners, respectively. When PLIN5 was increased in mouse β-cells in vivo, glucose tolerance after an acute exenatide challenge was improved. Therefore, the elevation of islet PLIN5 during fasting allows partitioning of FA into LD that is released upon refeeding to support PP insulin secretion in cAMP- and GPR40-dependent manners.
© 2015 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

Long chain fatty acids (LCFAs), which are ligands for the G-protein coupled receptor FFAR1 (GPR40), are increased in cow plasma after parturition, a period in which they are highly susceptible to infectious diseases. This study identified and analyzed the functional role of the FFAR1 receptor in bovine neutrophils, the first line of host defense against infectious agents. We cloned the putative FFAR1 receptor from bovine neutrophils and analyzed the sequence to construct a homology model. Our results revealed that the sequence of bovine FFAR1 shares 84% identity with human FFAR1 and 31% with human FFAR3/GPR41. Therefore, we constructed a homology model of bovine FFAR1 using human as the template. Expression of the bovine FFAR1 receptor in Chinese hamster ovary (CHO)-K1 cells increased the levels of intracellular calcium induced by the LCFAs, oleic acid (OA) and linoleic acid (LA); no increase in calcium mobilization was observed in the presence of the short chain fatty acid propionic acid. Additionally, the synthetic agonist GW9508 increased intracellular calcium in CHO-K1/bFFAR1 cells. OA and LA increased intracellular calcium in bovine neutrophils. Furthermore, GW1100 (antagonist of FFAR1) and U73122 (phospholipase C (PLC) inhibitor) reduced FFAR1 ligand-induced intracellular calcium in CHO-K1/bFFAR1 cells and neutrophils. Additionally, inhibition of FFAR1, PLC and PKC reduced the FFAR1 ligand-induced release of matrix metalloproteinase (MMP)-9 granules and reactive oxygen species (ROS) production. Thus, we identified the bovine FFAR1 receptor and demonstrate a functional role for this receptor in neutrophils activated with oleic or linoleic acid.

The G-protein-coupled receptor 40 (GPR40) is suggested to function as a transmembrane receptor for medium- to long-chain free fatty acids and is implicated to play a role in free fatty acids-mediated enhancement of glucose-stimulated insulin secretion from pancreas. However, the functional role of GPR40 in nervous system including somatosensory pain signaling has not been fully examined yet.
Intrathecal injection of GPR40 agonist (MEDICA16 or GW9508) dose-dependently reduced ipsilateral mechanical allodynia in CFA and SNL models and thermal hyperalgesia in carrageenan model. These anti-allodynic and anti-hyperalgesic effects were almost completely reversed by a GPR40 antagonist, GW1100. Immunohistochemical analysis revealed that GPR40 is expressed in spinal dorsal horn and dorsal root ganglion neurons, and immunoblot analysis showed that carrageenan or CFA inflammation or spinal nerve injury resulted in increased expression of GPR40 in these areas. Patch-clamp recordings from spinal cord slices exhibited that bath-application of either MEDICA16 or GW9508 significantly decreased the frequency of spontaneous excitatory postsynaptic currents in the substantia gelatinosa neurons of the three pain models.
Our results indicate that GPR40 signaling pathway plays an important suppressive role in spinal nociceptive processing after inflammation or nerve injury, and that GPR40 agonists might serve as a new class of analgesics for treating inflammatory and neuropathic pain.

GPR40 has been reported to be activated by long-chain fatty acids, such as docosahexaenoic acid (DHA). However, reports studying functional role of GPR40 in the brain are lacking. The present study focused on the relationship between pain regulation and GPR40, investigating the functional roles of hypothalamic GPR40 during chronic pain caused using a complete Freund's adjuvant (CFA)-induced inflammatory chronic pain mouse model. GPR40 protein expression in the hypothalamus was transiently increased at day 7, but not at days 1, 3 and 14, after CFA injection. GPR40 was co-localized with NeuN, a neuron marker, but not with glial fibrillary acidic protein (GFAP), an astrocyte marker. At day 1 after CFA injection, GFAP protein expression was markedly increased in the hypothalamus. These increases were significantly inhibited by the intracerebroventricular injection of flavopiridol (15 nmol), a cyclin-dependent kinase inhibitor, depending on the decreases in both the increment of GPR40 protein expression and the induction of mechanical allodynia and thermal hyperalgesia at day 7 after CFA injection. Furthermore, the level of DHA in the hypothalamus tissue was significantly increased in a flavopiridol reversible manner at day 1, but not at day 7, after CFA injection. The intracerebroventricular injection of DHA (50 µg) and GW9508 (1.0 µg), a GPR40-selective agonist, significantly reduced mechanical allodynia and thermal hyperalgesia at day 7, but not at day 1, after CFA injection. These effects were inhibited by intracerebroventricular pretreatment with GW1100 (10 µg), a GPR40 antagonist. The protein expression of GPR40 was colocalized with that of β-endorphin and proopiomelanocortin, and a single intracerebroventricular injection of GW9508 (1.0 µg) significantly increased the number of neurons double-stained for c-Fos and proopiomelanocortin in the arcuate nucleus of the hypothalamus. Our findings suggest that hypothalamic GPR40 activated by free long chain fatty acids might have an important role in this pain control system.