Neural tripeptide amide L-pyroglutamyl-L-histidyl-L-prolineamide (L-PHP, Thyrotropin-releasing hormone, TRH) is a fine molecular peptide that was first identified in the Central Nervous System (CNS) and discovered in many other regions of body later as a neuropeptide hormone or neuromodulator . L-PHP stimulates the thyroid stimulating hormone (TSH) after it is released from the hypothalamic nerve in the median eminence. L-PHP was named as its functional action-TRH . Beyond neuronal tissue, expression of L-PHP was also found in the pancreatic islets where it identifies to the Langerhans-insulin-producing beta cells . However, L-PHP expression and production is significantly different from its production in the nervous system; it is primarily expressed during the early developmental period in rat  and human fetal pancreatic tissue . L-PHP stimulates glucagon release and inhibits other pancreatic secretion other than TSH . In this review, based on evidence found in L-PHP gene knockout animal models and its function in regulating insulin release in pancreatic tissue , L-PHP may play an important role in carbohydrate metabolism and pancreatic L-PHP disruption may lead to the development of diabetes mellitus.
Expression of L-PHP in the pancreas: L-PHP is expressed in the insulin granules of β cells in pancreatic islets,  with high levels during the neonatal period but significantly decreased as postnatal development progresses . A Comparison with L-PHP expression, in the primary transition period between E12 and E14, shows insulin secretion in both rat and mouse while L-PHP remains unexpressed[4, 8]. During this period, insulin stained cells do not express any Rab3A, SNAP-25 (two molecules important for the control of insulin secretion) nor Glut 2 and granules resemble β cells. However, at E16, L-PHP expression was found and thereafter, high expression of molecules such as Glut2 and Pdx-1, which are necessary for insulin production, maturation and full insulin cell function, were found in the insulin and L-PHP positive cells. L-PHP’s significant expression coincides with factors for insulin production, maturation and insulin cell development suggesting that L-PHP is critical for insulin cells as they become functionally mature during early development.
2. Effects of L-PHP on pancreatic insulin secretion
Beyond regulating TSH, L-PHP is also found to be involved in the regulation of neuronal growth , facilitating spinal cord injury recovery , appetite control , and alcohol consumption . The most important role of L-PHP is considered to be its regulation of blood glucose levels
Pancreatic L-PHP can stimulate pancreatic endocrine function and/or endocrine cell development. The mechanisms as to how L-PHP regulates pancreatic β-cell development have not been identified. Gathering evidence from
L-PHP protects pancreatic tissue from damage and toxins like the reduction of glycodeoxycholic acid. Evidence suggests that L-PHP plays a critical role in β-cell maturation. During the phase of pancreatic development, which includes high levels of L-PHP expression in early pancreatic β-cell development, dexamethasone treatment eliminated the L-PHP peak and resulted in retarded β-cell development . Also, newborn rats were found to have reduced L-PHP levels due to maternal diabetes caused by streptozotocin (STZ) injection . The observation of ten-fold lower L-PHP in pups of diabetic rat followed by a postnatal day 5 elevation of L-PHP reducing blood glucose levels  suggest that L-PHP expression during β-cell development is important and it may prevent diabetes from developing in later life.
The L-PHP receptor consists of two major sub-types (R1 and R2, recently identified third type). Using RT-PCR, receptor R1 is identified as expressed in HIT-T15 (HIT) cells, a hamster clonal ß-cell line , and mouse pancreatic islets, but expression of R2 is not found. R2 was identified as expressed predominantly in the CNS, but not other tissue. By northern blot analysis it was found that R1 in pancreas is of 3.7-kb size and shares 93.3% homology with that in the pituitary. Evaluation of R1 function by receptor affinity found various kDa values in ß –cells . ß –cell intracellular calcium concentration was significantly increased by L-PHP and removal of extracellular calcium does not change this effect . Our group work has shown R1 expression in rat-derived β-cell lines as well as whole pancreas that included nonislet tissue . R1 receptor was also found to associate with EGF receptor function called cross linking  (Fig. 2).
3. Regulation of L-PHP in the pancreas
4. L-PHP alteration of gene expression modifys microenvironment within the pancreas
Pancreatic microenvironment alteration by L-PHP has been reported . The findings show that multiple functional genes in rat pancreas were influenced by L-PHP
5. Regulation of β-cell proliferation by signal pathways from L-PHP to growth hormone activity in pancreatic islet
L-PHP has been reported to stimulate R1 and dissociate the GPCR complex, activating protein kinase C  and mitogen-activated protein kinase (MAPK)  in both a PKC-dependent and a PKC-independent manner in the neuronal cell lines . These effects may involve activation of tyrosine kinase, which leads to the activation of Ras and MAPK cascade. The signaling pathways initiating from G-coupled L-PHP receptor in activating MAPK may overlap with the receptor tyrosine kinases activating the Ras-MAPK cascade [31, 32]. There is evidence that L-PHP and EGF have overlapping activities  leading to the stimulation of tyrosine phosphorylation of EGF receptors in GH3 cells, a pituitary cell line . L-PHP-induced EGF receptor phosphorylation led to the recruitment of adapter protein Grb2 and Shc in GH3 cells. The hypothesis that L-PHP would activate EGF receptors in β cells through multiple pathways is tested, and data indicated that L-PHP trans-activates EGF receptors through several intra-and extracellular pathways, which are distinguished from pituitary-derived cell lines. R1 can initiate multiple signal transduction pathways to activate the epidermal growth factor (EGF) receptor in pancreatic β cells . By initiating R1 G-protein-coupled receptor (GPCR) and dissociated αβγ complex, L-PHP (200nM) activates tyrosine residues at Tyr845, (a known target for Src) and Tyr1068 in the EGF receptor complex in an immortalized mouse β-cell line, βTC-6. Through manipulating the activation of Src, PKC and heparin-binding EGF-like growth factor (HB-EGF) with corresponding individual inhibitors and activators, multiple signal transduction pathways linking L-PHP to EGF receptors in βTC-6 cell lines have been revealed. The pathways include the activation of Src kinase and the release of heparin-binding EGF as a consequence of MMP3 activation. Alternatively, L-PHP inhibited PKC activity by reducing EGF receptor serine/threonine phosphorylation, thereby enhancing tyrosine phosphorylation. L-PHP receptor activation of Src may have a central role in mediating the effects of L-PHP on the EGF receptor (Fig. 3). The activation of the EGF receptor by L-PHP in multiple circumstances may have important implications for pancreatic β cell biology. Since EGF receptor expression has been found to have a high activity during the embryonic developmental period [4, 36], the possibility exists that L-PHP activation of EGF receptors in pancreatic β cells may play a role in β-cell development.
The small sized L-PHP neuropeptide may play a significant role in direct regulation of pancreatic β-cell function and, through modulation of pancreatic microenvironment, support β-cell survival. The role of L-PHP may be similar to that of the gut peptide GLP-1, that increases β-cell regeneration, but may also have a role in inducing adult stem cell differentiation into functional β-cells during pancreatic tissue injury, which may be significant for diabetic therapy.
7. Future directions
Rat islet cell function can be recovered 90-95% from a pancreatectomy after application of glucagon-like peptide 1(GLP-1) . This β-cell regeneration from damaged rat pancreas has also been mimicked by STZ damaged rat pancreas following administration of L-PHP . However, human islet β-cell regeneration may differ from rat and it may require a totally different microenvironment. In order to initiate human islet β-cell functional recovery from damage or loss, pancreatic stem cells or stem cells from other tissue, such as bone marrow, must be able to in vivo differentiate into multiple types of endocrine cells (αβγ) to reconstitute a new endocrine system in response to glucose challenge. Initiating L-PHP generation
This work was partially supported by Roger Williams Hospital Research fund, and NIH Grant Number 1R01DK097380-01A1 from National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) and National Institute of General Medical Sciences (NIGMS). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH.
Engler D, Scanlon MF, Jackson IM. Thyrotropin-releasing hormone in the systemic circulation of the neonatal rat is derived from the pancreas and other extraneural tissues. J Clin Invest 1981; 67: 800-8.
Reichlin S, Saperstein R, Jackson IM, Boyd AE 3rd, Patel Y. Hypothalamic hormones. Annu Rev Physiol 1976; 38: 389-424.
Wu P, Jackson IM. Identification, characterization and localization of thyrotropin-releasing hormone precursor peptides in perinatal rat pancreas. Regul Pept 1988; 22(4): 347-60.
Basmaciogullari A, Cras-Meneur C, Czernichow P, Scharfmann R. Pancreatic pattern of expression of thyrotropin-releasing hormone during rat embryonic development. J Endocrinol 2000; 166: 481-8.
Yamagishi K. Pancreatic exocrine and endocrine functions stimulated with secretin and thyrotropin-releasing hormone in patients with hyperparathyroidism. Nihon Geka Gakkai Zasshi 1992; 93(5): 494-504.
Yamada M, Saga Y, Shibusawa N, et al. Tertiary hypothyroidism and hyperglycemia in mice with targeted disruption of the thyrotropin-releasing hormone gene. Proc Natl Acad Sci USA 1997; 94: 10862-7.
Fragner P, Ladram A, Aratan de Leon S. Triiodothyronine down-regulates thyrotropin-releasing hormone (TRH) synthesis and decreases pTRH-(160-169) and insulin releases from fetal rat islets in culture. Endocrinology 1999; 140(9): 4113-9.
Yamaoka T, Itakura M. Development of pancreatic islets (review). Int J Mol Med 1999; 3: 247-61.
Koenig ML, Sgarlat CM, Yourick DL, Long JB, Meyerhoff JL. In vitroneuroprotection against glutamate-induced toxicity by pGlu-Glu-Pro-NH(2) (EEP). Peptides 2001; 22: 2091-7.
Behrmann DL, Bresnahan JC, Beattie MS. Modeling of acute spinal cord injury in the rat: neuroprotection and enhanced recovery with methylprednisolone, U-74006F and YM-14673. Exp Neurol. 1994;126: 61-75.
Stanley SA, Small CJ, Murphy KG, et al. Actions of cocaine-and amphetamine-regulated transcript (CART) peptide on regulation of appetite and hypothalamo-pituitary axes in vitroand in vivoin male rats Brain Res. 2001; 893: 186-94.
de Gortari P, Méndez M, Rodríguez-Keller I, Pérez-Martínez L, Joseph-Bravob P. Acute ethanol administration induces changes in TRH and proenkephalin expression in hypothalamic and limbic regions of rat brain. Neurochem Int. 2000; 37: 483-96.
Amir S, Jackson IM. Immunological blockade of endogenous thyrotropin-releasing hormone impairs recovery from hyperglycemia in mice. Brain Res. 1988; 462: 160-2.
Chen Y, Uemura K, Yoshioka S, et al. Centrally administered TRH-induced insulin secretion is impaired in the Otsuka-Long-Evans-Tokushima Fatty rats, a model of spontaneous non-insulin-dependent diabetes mellitus. J Auton Nerv Syst 1998; 71: 10-7.
Rondeel JM, de Greef WJ, Heide R, Visser TJ. Hypothalamo-hypophysial-thyroid axis in streptozotocin-induced diabetes. Endocrinology 1992;130: 216-20.
Ishiguro T, Iguchi A, Kunoh Y, et al. Relative contribution of nervous system and hormones to hyperglycemia induced by thyrotropin-releasing hormone in fed rats. Neuroendocrinology 1991; 54: 1-6.
Amir S. Thyrotropin-releasing hormone (TRH) blocks glucagon-induced hyperglycemia in mice: dissociation of the antihyperglycemic and pituitary actions of TRH. Brain Res 1988; 455: 201-3.
Roper MG, Qian WJ, Zhang BB, Kulkarni RN, Kahn CR, Kennedy RT. Effect of the insulin mimetic L-783,281 on intracellular Ca2+and insulin secretion from pancreatic beta-cells. Diabetes 2002; 51: S43-9.
Maltese JY, Giraud P, Kowalski C, et al. Ontogenetic expression of peptidyl-glycine alpha-amidating monooxygenase mRNA in the rat pancreas. Biochem Biophys Res Commun 1989; 158: 244-50.
Vara E, Idahl LA, Lindström P, Sehlin J, Tamarit-Rodriguez J. Insulin, glucagon, somatostatin, and thyrotropin-releasing hormone content and secretion by perifused fetal rat islets during culture. Acta Endocrinol (Copenh) 1990; 123: 353-8.
Glasbrenner B, Malfertheiner P, Duntas L, Büchler M, Bereiter T, Ditschuneit H. Effects of TRH on pancreatic growth and secretion in rats. Pancreas 1990; 5: 37-41.
Strbák V, Ouafik LH, Resetková E, et al.Thyrotropin releasing hormone in the pancreas of newborn rats from streptozotocin-treated mothers. Life Sci 1989; 44: 779-87.
Yamada M, Shibusawa N, Hashida T, et al. Expression of thyrotropin-releasing hormone (TRH) receptor subtype 1 in mouse pancreatic islets and HIT-T15, an insulin-secreting clonal beta cell line. Life Sci 2000; 66:1119-25.
Alhan E, Küçüktülü U, Erçin C, Efe H, Al S. The effects of calcium channel blocker and thyrotropin releasing hormone on acute necrotizing pancreatitis in rats. Res Exp Med (Berl) 1999; 199: 51-8.
Luo LG, Yano N. Expression of thyrotropin-releasing hormone receptor in immortalized beta-cell lines and rat pancreas. J Endocrinol 2004; 181: 401-12.
Doong ML, Yang H. Intravenous glucose infusion decreases intracisternal thyrotropin-releasing hormone induced vagal stimulation of gastric acid secretion in anesthetized rats. Neurosci Lett 2003; 340: 49-52.
Pizzi M, Boroni F, Benarese M, Moraitis C, Memo M, Spano P. Neuroprotective effect of thyrotropin-releasing hormone against excitatory amino acid-induced cell death in hippocampal slices. Eur J Pharmacol 1999; 370: 133-7.
Yano N, Luo L. Effect of thyrotropin releasing hormone (TRH) on gene expressions in rat pancreas: approach by microarray hybridization. JOP 2004; 5: 193-204.
Buteau J, Foisy S, Rhodes CJ, Carpenter L, Biden TJ, Prentki M. Protein kinase Czeta activation mediates glucagon-like peptide-1-induced pancreatic beta-cell proliferation. Diabetes 2001; 50: 2237-43.
Smith J, Yu R, Hinkle PM. Activation of MAPK by TRH requires clathrin-dependent endocytosis and PKC but not receptor interaction with beta-arrestin or receptor endocytosis. Mol Endocrinol 2001; 15:1539-48.
Palomero T, Barros F, del Camino D, Viloria CG, de la Peña P. A G protein beta gamma dimer-mediated pathway contributes to mitogen-activated protein kinase activation by thyrotropin-releasing hormone receptors in transfected COS-7 cells. Mol Pharmacol 1998; 53: 613-22.
Ohmichi M, Sawada T, Kanda Y, et al. Thyrotropin-releasing hormone stimulates MAP kinase activity in GH3 cells by divergent pathways. Evidence of a role for early tyrosine phosphorylation. J Biol Chem 1994; 269: 3783-8.
Andreev J, Galisteo ML, Kranenburg O, et al. Src and Pyk2 mediate G-protein-coupled receptor activation of epidermal growth factor receptor (EGFR) but are not required for coupling to the mitogen-activated protein (MAP) kinase signaling cascade. J Biol Chem 2001; 276: 20130-5.
Wang YH, Jue SF, Maurer RA. Thyrotropin-releasing hormone stimulates phosphorylation of the epidermal growth factor receptor in GH3 pituitary cells. Mol Endocrinol. 2000; 14: 1328-37.
Luo L, Yano N, Luo JZ. The molecular mechanism of EGF receptor activation in pancreatic beta-cells by thyrotropin-releasing hormone. Am J Physiol Endocrinol Metab 2006; 290: E889-99.
Cras-Méneur C, Elghazi L, Czernichow P, Scharfmann R. Epidermal growth factor increases undifferentiated pancreatic embryonic cells in vitro: a balance between proliferation and differentiation. Diabetes 2001; 50: 1571-9.
Luo LG. International Society for stem cell research (ISSCR) 2nd Annual Meeting, Boston MA 2004; Abstract 183:147.
Li Y, Hansotia T, Yusta B, Ris F, Halban PA, Drucker DJ. Glucagon-like peptide-1 receptor signaling modulates beta cell apoptosis. J Biol Chem 2003; 278: 471-8.
Luo LG, Yano N, Jackson I. 86th Annual Endocrine Meeting 2004 New Orleans Louisana June 16-19. 2004; Abstract: OR 46-2, pp 141.