Different responses in feed intake have been noticed followi

Different responses in feed intake have been noticed following administration of exogenous GHRL (peripherally or centrally) depending on animal species. In mammals, injection of GHRL stimulates food intake (orexigenic effect) (reviewed in Kaiya et al., 2013b). In contrast, in domestic birds, exogenous GHRL reduces appetite (anorexigenic effect) (Geelissen et al., 2006, Lotfi et al., 2013). In support to the anorexigenic role of GHRL in birds, GHRL treatment inhibits the hypothalamic fuel sensor AMP-kinase (AMPK) (Xu et al., 2011) which is opposite to the stimulatory effect of GHRL on the hypothalamic AMPK in mammals (Stark et al., 2013). Central GHRL injection also decreases gamma aminobutyric Pemetrexed mg (GABA) synthesis in chick brain, which suppresses the appetite (Jonaidi et al., 2012). The anorexigenic effect of GHRL depends on species, strain, rearing conditions, age, bodyweight and route of GHRL administration (Kaiya et al., 2013b). On the other hand, mammalian GHRL is a potent stimulator of growth hormone (GH) release and enhances feeding and weight gain to regulate energy balance. Although mammalian and avian species differ in their regulation of GH secretion, some studies have demonstrated in vivo GH responses to GHRL in chickens, as in mammals. GHRL has been shown to increase plasma GH concentrations in immature chickens (Ahmed and Harvey, 2002, Baudet and Harvey, 2003, Kaiya et al., 2002). In addition, it was suggested that endogenous chicken GHRL is a “hunger signal” which is similar to mammalian GHRL (Kaiya et al., 2013b, Kaiya et al., 2008), however the authors failed to prove their hypothesis due to lack of studies on association between polymorphisms in endogenous GHRL (and/or its receptor) and expression of GHRL. In chicken, 37 single nucleotides polymorphisms (SNPs) and a ‘GGTACA’ indel were detected in the full length of GHSR gene (Nie et al., 2005). Only few number of these SNPs, which are mainly located in exon 2, significantly associated with growth traits (Fang et al., 2010, Lei et al., 2007). Recently, another SNP (C3286>T) was also detected in exon 2 (at nucleotide number 74), and association analysis showed a significant association (P<0.01) with growth traits (daily feed intake, and average daily gain) and body composition traits (abdominal fat weight, AFW and %AFW) (Darzi Niarami et al., 2014). This suggests that most valuable SNPs of chicken GHSR are located in exon 2. However, these previous association studies did not investigate the effect of these SNPs on the expression and/or level GHRL and how this can affect appetite. In a trial to find a suitable answer for the question “Is really endogenous GHRL a hunger signal in chickens?”, we searched for polymorphisms in GHSR exon 2 and studied their associations with feed intake and growth traits in addition to gene expression and serum level of GH and GHRL in a local Egyptian breed, Mandara.
Material and methods
Results
Discussion In the present study, two adjacent SNPs, A239G (led to synonymous mutation) and G244A (led to non-synonymous mutation which changed lysine into arginine), were detected in exon 2 of the GHSR gene of Mandara chicken. None of these SNPs was found in the pure Egyptian breeds, Fayoumy and Dandarawy (personal observation, unpublished data), or foreign breeds studied so far (Darzi Niarami et al., 2014, Fang et al., 2010, Lei et al., 2007, Nie et al., 2005). Mandara breed is not a pure Egyptian breed and is produced by crossing between Dokki-4 females and Alexandria males, which are produced by crossing between pure Egyptian breed (Fayoumy) and foreign breeds (Barred Plymouth Rock, Rhode Island Red, and White Leg Horn) (Abd El-Gawad et al., 1983). Fayoumy breeds have lower growth rate and meat production than foreign breeds. The hybrid chickens Mandara have higher body weight, growth rate and meat production than Fayoumy chickens. Therefore, it is possible that A239G and G244A SNPs in Mandara chicken produced by this crossing and these two very close SNPs (only 4bp distance) may be associated with chicken growth performance. As expected from their close positions, the two SNPs were in a very strong linkage disequilibrium (r2=0.914; D′=1.0), meaning that these SNPs are co-inherited (dependent) and therefore their effect on the growth traits may be mutual. This prompts us to study the association of the combined genotypes (haplotypes) of both SNPs instead of individual SNP. This combined genotypes produced only three haplotypes (GG/GG, GG/AG, AG/AG), which associated significantly with feed intake, growth traits (body weight, ADG, shank length, keel length, chest circumference) in Mandara chicken at age from 4 to 16w. Chickens with the homozygous GG/GG haplotype showed higher growth performance and so they could be selected to obtain superior growth performance at growing stage from 4 to 16w. In agreement, Fang et al. (2010) reported that combined genotypes (haplotypes) for GHSR SNPs appeared advantageous for growth and carcass traits in the breeding program of Chinese chicken. Some other SNPs were detected in poultry GHSR gene: C3286T, T640C and C1459T in chicken and A3427T in duck and their significant associations with growth and carcass traits were separately studied (Darzi Niarami et al., 2014, Nie et al., 2009).