Postmortem mitochondria function in longissimus lumborum of Angus and Brahman steers.

Mitochondria function and integrity may impact postmortem metabolism and meat quality development. Adaptations in heat tolerant Brahman may persist to limit cellular stress postmortem. Our objective was to evaluate glycolysis, pH decline, and mitochondria function in longissimus lumborum (LL) from Angus and Brahman steers (N = 28) early postmortem (1 to 6 h) and after rigor (24 h). We evaluated metabolites of anaerobic glycolysis, ATP, pH, and temperature, and determined mitochondria oxygen consumption rate (OCR) in permeabilized fibers. The main effects of breed (b) and time (t) and the interaction were tested. Brahman LL contained greater ATP during the first 6 h postmortem; Brahman also tended to exhibit a slower pH decline (b × t, P = 0.07) and more rapid temperature decline (b × t, P < 0.001), but metabolites of anaerobic glycolysis were not different. Mitochondria in Brahman and Angus LL were well-coupled and respired at 1 h postmortem. However, outer membrane integrity became increasingly compromised postmortem (t, P < 0.001). Brahman tended to exhibit greater electron transport system capacity (b, P < 0.1) and had greater capacity for oxidative phosphorylation (complex I and II substrates) at 6 h compared with Angus (P < 0.001). In totality, greater ATP, slower pH decline, and enhanced mitochondria capacity indicate that Brahman possess mitochondrial properties or cellular adaptations that help protect the cell during energy stress postmortem. Slower pH and more rapid temperature decline in LL from Brahman may also help preserve mitochondria function postmortem.

National Beef Quality Audit-2022: Transportation, mobility, live cattle, and hide assessments to determine producer-related defects that affect animal welfare and the value of market cows and bulls at processing facilities.

The National Beef Quality Audit (NBQA)-2022 serves as a benchmark of the current market cow and bull sectors of the U.S. beef industry and allows comparison to previous audits as a method of monitoring industry progress. From September 2021 through May 2022, livestock trailers (n = 125), live animals (n = 5,430), and post-slaughter hide-on animals (n = 6,674) were surveyed at 20 commercial beef processing facilities across the U.S. Cattle were transported in a variety of trailer types for an average distance of 490.6 km and a mean transport time of 6.3 h. During transit, cattle averaged 2.3 m2 of trailer space per animal indicating sufficient space was provided according to industry guidelines. Of all trailers surveyed, 55.3% transported cattle from an auction barn to a processing facility. When surveyed, 63.6% of all truck drivers reported to be Beef Quality Assurance certified. The majority (77.0%) of cattle were sound when evaluated for mobility. Mean body condition scores (9-point scale) for beef cows and bulls were 3.8 and 4.4, respectively, whereas mean body condition scores (5-point scale) for dairy cows and bulls were 2.3 and 2.6, respectively. Of the cattle surveyed, 45.1% had no visible live animal defects, and 37.9% had only a single defect. Of defects present in cows, 64.6% were attributed to an udder problem. Full udders were observed in 47.5% of all cows. Nearly all cattle were free of visible abscesses and knots (97.9% and 98.2%, respectively). No horns were observed in 89.4% of all cattle surveyed. Beef cattle were predominantly black-hided (68.9% and 67.4% of cows and bulls, respectively). Holstein was the predominant dairy animal observed and accounted for 85.7% of the cows and 98.0% of the bulls. Only 3.1% of all animals had no form of identification. Findings from the NBQA-2022 show improvements within the industry and identify areas that require continued education and research to improve market cow and bull welfare and beef quality.

Presence of KREMEN receptors for DKK1 in the preimplantation bovine embryo.

The WNT inhibitory protein DKK1 has been shown to regulate development of the preimplantation embryo to the blastocyst stage. In cattle, DKK1 increases the number of trophectoderm cells that are the precursor of the placenta. DKK1 can affect cells by blocking WNT signaling through its receptors KREMEN1 and KREMEN2. Here it was shown that the mRNA for KREMEN1 and KREMEN2 decline as the embryo advances in development. Nonetheless, immunoreactive KREMEN1 was identified in blastocysts using Western blotting. DKK1 also decreased amount of immunoreactive CTNNB1 in blastocysts, as would be expected if DKK1 was signaling through a KREMEN-mediated pathway. Thus, it is likely that KREMEN1 functions as a receptor for DKK1 in the preimplantation bovine embryo.

Actions of WNT family member 5A to regulate characteristics of development of the bovine preimplantation embryo†.

WNT signaling is important for regulation of embryonic development. The most abundant WNT gene expressed in the bovine endometrium during the preimplantation period is WNT5A. One objective was to determine whether WNT5A regulates competence of the bovine preimplantation embryo to become a blastocyst and alters the number of cells in the inner cell mass and trophectoderm. A second objective was to delineate features of the cell-signaling mechanisms involved in WNT5A actions. WNT5A caused a concentration-dependent increase in the proportion of embryos developing to the blastocyst stage and in the number of inner cell mass cells in the resultant blastocysts. A concentration of 200 ng/mL was most effective, and a higher concentration of 400 ng/mL was not stimulatory. Bovine serum albumin in culture reduced the magnitude of effects of WNT5A on development to the blastocyst stage. WNT5A affected expression of 173 genes at the morula stage; all were upregulated by WNT5A. Many of the upregulated genes were associated with cell signaling. Actions of WNT5A on development to the blastocyst stage were suppressed by a Rho-associated coiled-coil kinase (ROCK) signaling inhibitor, suggesting that WNT5A acts through Ras homology gene family member A (RhoA)/ROCK signaling. Other experiments indicated that actions of WNT5A are independent of the canonical ß-catenin signaling pathway and RAC1/c-Jun N-terminal kinase (JNK) signaling. This is the first report outlining the actions of WNT5A to alter the development of the mammalian embryo. These findings provide insights into how embryokines regulate maternal-embryonic communication.

Connecting Heat Tolerance and Tenderness in Bos indicus Influenced Cattle.

Bos indicus cattle are widely utilized in tropical and subtropical climates. Their heat tolerance and parasite resistance are integral for beef production in these regions; however, a reputation for excitable temperaments, slower growth, and variation in tenderness has limited their use in commercial beef production. This suggests that there is antagonism between heat tolerance and meat production traits. Meat quality characteristics are determined by the properties of skeletal muscle as well as conditions during slaughter and processing. Thus, it is possible that adaptations related to heat tolerance in the living animal affect tenderness and other meat quality attributes. Since muscle represents a large proportion of body mass, relatively small changes at the cellular level could impact overall heat production of the animal. Specifically, protein degradation and mitochondria function are aspects of organ and cellular metabolism that may help limit heat production and also have a connection to tenderness. Protein degradation postmortem is critical to structural changes that enhance tenderness whereas mitochondria may influence tenderness through their roles in energy metabolism, calcium regulation, cell death signaling, and oxidative stress. This review explores potential relationships between cellular metabolism in vivo and beef quality development in Bos indicus and Bos indicus influenced cattle.

Mitochondrial oxygen consumption in early postmortem permeabilized skeletal muscle fibers is influenced by cattle breed.

Functional properties and integrity of skeletal muscle mitochondria (mt) during the early postmortem period may influence energy metabolism and pH decline, thereby impacting meat quality development. Angus typically produce more tender beef than Brahman, a Bos indicus breed known for heat tolerance. Thus, our objectives were to compare mt respiratory function in muscle collected early postmortem (1 h) from Angus and Brahman steers (n = 26); and to evaluate the effect of normal and elevated temperature on mt function ex vivo. We measured mt oxygen consumption rate (OCR) in fresh-permeabilized muscle fibers from Longissimus lumborum (LL) at 2 temperatures (38.5 and 40.0 °C) and determined citrate synthase (CS) activity and expression of several mt proteins. The main effects of breed, temperature, and their interaction were tested for mt respiration, and breed effect was tested for CS activity and protein expression. Breed, but not temperature (P > 0.40), influenced mt OCR (per tissue weight), with Brahman exhibiting greater complex I+II-mediated oxidative phosphorylation capacity (P = 0.05). Complex I- and complex II-mediated OCR also tended to be greater in Brahman (P = 0.07 and P = 0.09, respectively). Activity of CS was higher in LL from Brahman compared to Angus (P = 0.05). Expression of specific mt proteins did not differ between breeds, except for higher expression of adenosine triphosphate (ATP) synthase subunit 5 alpha in Brahman muscle (P = 0.04). Coupling control ratio differed between breeds (P = 0.05), revealing greater coupling between oxygen consumption and phosphorylation in Brahman. Our data demonstrate that both Angus and Brahman mt retained functional capacity and integrity 1-h postmortem; greater oxidative phosphorylation capacity and coupling in Brahman mt could be related to heat tolerance and impact early postmortem metabolism.

Inhibition of mitochondrial calcium uniporter enhances postmortem proteolysis and tenderness in beef cattle.

The purpose of this study was to examine the role of mitochondria in postmortem calcium homeostasis and its effect on proteolysis and tenderness. We hypothesized that mitochondria buffer cytosolic calcium levels and delay the activation of calpain-1 and subsequently the development of meat tenderness. To test this hypothesis, pre-rigor bovine longissimus thoracis et lumborum muscle samples were injected with DS16570511 to inhibit mitochondrial calcium uptake. Free calcium, tenderness, texture profile analysis (TPA), calpain-1 activity, and proteolysis were evaluated over a 336 h aging period. Inhibition of mitochondrial calcium uptake increased (P < .0001) cytosolic calcium concentration and calpain-1 autolysis and activity at 24 h compared to control steaks. Further, tenderness and TPA at 168 and 336 h, calpastatin degradation at 24 h, and proteolysis at 168 h were all enhanced (P < .05) in the treated steaks. Collectively, these data indicate that inhibition of mitochondrial calcium uptake can enhance postmortem proteolysis and tenderization through an early activation of calpain-1.

Resistance to pH decline and slower calpain-1 autolysis are associated with higher energy availability early postmortem in Bos taurus indicus cattle.

Beef from Bos taurus indicus is associated with toughness compared to Bos taurus taurus, suggesting there is antagonism between adaptability to heat and beef quality. Resistance to cellular stress in muscle may be protective postmortem, thereby delaying its conversion to meat. Therefore, our objective was to determine pH decline, calpain-1 and caspase 3 activation, and proteolysis in different biological cattle types. Angus, Brangus, and Brahman steers (n = 18) were harvested, and Longissimus lumborum were assessed postmortem for pH decline, ATP content, protease activation, and calpastatin content; and myofibrillar protein degradation was evaluated in beef aged to 14d. Brahman Longissimus lumborum exhibited resistance to pH decline, greater ATP content at 1 h, and delayed calpain-1 autolysis. Although content of caspase-3 zymogen was lower in Brahman, there was no evidence of caspase-3 mediated proteolysis. Greater resistance to energetic and pH changes early postmortem in Brahman Longissimus lumborum are associated with calpain-1 autolysis but not mitochondria mediated apoptosis.

Technical note: Protocol for electrophoretic separation of bovine myosin heavy chain isoforms and comparison to immunohistochemistry analysis.

Myosin heavy chain (MyHC) isoform composition is a primary determinant of contractile speed of muscle fibers. Currently, bovine MyHC isoforms are evaluated using time-consuming histochemical analysis by immunflourescence or ATPase activity. Electrophoretic separation of MyHC isoforms is more rapid; however, a reliable procedure without use of gradients has not been validated for cattle. Therefore, our objectives were to develop and validate a procedure for separating bovine MyHC isoforms (I, IIa, and IIx) using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and compare results to immunohistochemistry (IHC) analysis. Muscle samples were collected from masseter, sternomandibularis, diaphragm, longissimus lumborum, and cutaneous trunci within 1.5 h postmortem. To determine appropriate conditions for electrophoretic separation, several parameters of gel composition were varied. Bovine MyHC isoforms were clearly separated by increasing glycerol content of polyacrylamide gels to 37%. Identity of MyHC isoforms was confirmed using western blotting. Percent MyHC composition evaluated by gel electrophoresis was consistent with IHC (P > 0.2). Thus, SDS-PAGE produces clear separation of MyHC isoforms, and is a viable alternative to IHC-based methods.

Quantitative Proteomics and Phosphoproteomics Analysis Revealed Different Regulatory Mechanisms of Halothane and Rendement Napole Genes in Porcine Muscle Metabolism.

Pigs with the Halothane (HAL) or Rendement Napole (RN) gene mutations demonstrate abnormal muscle energy metabolism patterns and produce meat with poor quality, classified as pale, soft, and exudative (PSE) meat, but it is not well understood how HAL and RN mutations regulate glucose and energy metabolism in porcine muscle. To investigate the potential signaling pathways and phosphorylation events related to these mutations, muscle samples were collected from four genotypes of pigs, wild type, RN, HAL, and RN-HAL double mutations, and subjected to quantitative proteomic and phosphoproteomic analysis using the TiO2 enrichment strategy. The study led to the identification of 932 proteins from the nonmodified peptide fractions and 1885 phosphoproteins with 9619 phosphorylation sites from the enriched fractions. Among them, 128 proteins at total protein level and 323 phosphosites from 91 phosphoproteins were significantly regulated in mutant genotypes. The quantitative analysis revealed that the RN mutation mainly affected the protein expression abundance in muscle. Specifically, high expression was observed for proteins related to mitochondrial respiratory chain and energy metabolism, thereby enhancing the muscle oxidative capacity. The high content of UDP-glucose pyrophosphorylase 2 (UGP2) in RN mutant animals may contribute to high glycogen storage. However, the HAL mutation mainly contributes to the up-regulation of phosphorylation in proteins related to calcium signaling, muscle contraction, glycogen, glucose, and energy metabolism, and cellular stress. The increased phosphorylation of Ca2+/calmodulin-dependent protein kinase II (CAMK2) in HAL mutation may act as a key regulator in these processes of muscle. Our findings indicate the different regulatory mechanisms of RN and HAL mutations in relation to porcine muscle energy metabolism and meat quality.

Association of µ-Calpain and Calpastatin Polymorphisms with Meat Tenderness in a Brahman-Angus Population.

Autogenous proteolytic enzymes of the calpain family are implicated in myofibrillar protein degradation. As a result, the µ-calpain gene and its specific inhibitor, calpastatin, have been repeatedly investigated for their association with meat quality traits in cattle; however, no functional mutation has been identified for these two genes. The objectives of this study were: (1) to assess breed composition effect on tenderness; (2) to perform a linkage disequilibrium (LD) analysis in µ-calpain and calpastatin genes as well as an association analyses with tenderness; and (3) to analyze putative functional SNPs inside the significant LD block for an effect on tenderness. Tenderness measurements and genotypes for 16 SNPs in µ-calpain gene and 28 SNPs in calpastatin gene from 673 steers were analyzed. A bioinformatic analysis identified "putative functional SNPs" inside the associated LD block - polymorphisms able to produce a physical and/or chemical change in the DNA, mRNA, or translated protein in silico. Breed composition had a significant (P < 0.0001) effect on tenderness where animals with more than 80% Angus composition had the most tender meat. One 11-kb LD-block and three LD-blocks of 37, 17, and 14 kb in length were identified in the µ-calpain and calpastatin genes, respectively. Out of these, the LD-block 3 in calpastatin, tagged by SNPs located at 7-98566391 and 7-98581038, had a significant effect on tenderness with the TG-CG diplotype being approximately 1 kg more tender than the toughest diplotype, TG-CG. A total of 768 SNPs in the LD-block 3 of calpastatin were included in the bioinformatic analysis, and 28 markers were selected as putative functional SNPs inside the LD-block 3 of calpastatin; however, none of them were polymorphic in this population. Out of 15 initial polymorphisms segregating inside the LD-block 3 of calpastatin in this population, markers ARSUSMARC116, Cast5, rs730723459, and rs210861835 were found to be significantly associated with tenderness.

Presence of oxygen and mitochondria in skeletal muscle early postmortem.

Anaerobic glycolysis dominates energy metabolism postmortem. Even so, however, recent studies suggest mitochondria can modify postmortem energy metabolism and may contribute to pH decline, possibly affecting the transformation of muscle to meat and fresh meat quality development. Because oxygen is a necessary component of mitochondrial function, oxygenation of porcine and bovine longissimus thoracis et lumborum was determined postmortem using NIR spectroscopy. The ratio of oxy- to deoxymyoglobin decreased with time postmortem in both species. Metabolic analyses of muscle samples collected over the same timeframe also revealed fluctuations in TCA intermediates. Finally, mitochondria collected from muscle of electrically stimulated carcasses differed from those of non-stimulated muscle. Collectively, these data support the thesis that muscle mitochondria function early postmortem and may play a more active part in pH decline and possibly meat quality development.

Brahman genetics influence muscle fiber properties, protein degradation, and tenderness in an Angus-Brahman multibreed herd.

The objective of this study was to determine the influence of Brahman genetics on muscle contractile and metabolic phenotype and postmortem proteolysis. Cattle used in this study represent a continuous spectrum of Angus-Brahman genetic variation. Steers were harvested and Longissimus samples were collected at 1.5h, 24h, and 14d postmortem. Proteolysis during the 14d aging period was evaluated, along with Warner-Bratzler shear force (WBSF) and trained sensory panel tenderness. Myosin heavy chain composition and enzymatic activity were used to evaluate fiber type characteristics. As Brahman influence increased, WBSF increased and sensory tenderness decreased. Calpain-1 autolysis decreased as Brahman percentage increased, and corresponded with reduced degradation of troponin-T, desmin, and titin. Increasing Brahman percentage was associated with greater citrate synthase activity and greater cross-sectional area of type IIx fibers. Brahman-influenced cattle produced tougher steaks and exhibited decreased protein degradation. Thus, Brahman genetics impacted not only the calpain-calpastatin system, but also muscle fiber size and metabolic properties.

A mitochondrial protein increases glycolytic flux.

The purpose of this study was to determine the role of mitochondria in postmortem muscle metabolism. Isolated mitochondria were incorporated into a reaction buffer that mimics postmortem glycolysis with or without mitochondrial electron transport chain inhibitors. Addition of mitochondria lowered pH values at 240 and 1440min regardless of inhibitors. Reduction in pH was accompanied by enhanced glycogen degradation and lactate accumulation. To explore the mechanism responsible for this exaggerated metabolism, mitochondrial preparations were mechanically disrupted and centrifuged. Resulting supernatants and pellets each were added to the in vitro model. Mitochondrial supernatants produced similar effects as those including intact mitochondria. To narrow further our target of investigation, mitochondrial supernatants were deproteinized with perchloric acid. The effect of mitochondrial supernatant was lost after perchloric acid treatment. These data indicate that a mitochondrial-based protein is capable of increasing glycolytic flux in an in vitro model and may partially explain acid meat development in highly oxidative AMPKγ3R200Q mutated pigs.

Gain of function AMP-activated protein kinase γ3 mutation (AMPKγ3R200Q) in pig muscle increases glycogen storage regardless of AMPK activation.

Chronic activation of AMP-activated protein kinase (AMPK) increases glycogen content in skeletal muscle. Previously, we demonstrated that a mutation in the ryanodine receptor (RyR1(R615C)) blunts AMPK phosphorylation in longissimus muscle of pigs with a gain of function mutation in the AMPKγ3 subunit (AMPKγ3(R200Q)); this may decrease the glycogen storage capacity of AMPKγ3(R200Q) + RyR1(R615C) muscle. Therefore, our aim in this study was to utilize our pig model to understand how AMPKγ3(R200Q) and AMPK activation contribute to glycogen storage and metabolism in muscle. We selected and bred pigs in order to generate offspring with naturally occurring AMPKγ3(R200Q), RyR1(R615C), and AMPKγ3(R200Q) + RyR1(R615C) mutations, and also retained wild-type littermates (control). We assessed glycogen content and parameters of glycogen metabolism in longissimus muscle. Regardless of RyR1(R615C), AMPKγ3(R200Q) increased the glycogen content by approximately 70%. Activity of glycogen synthase (GS) without the allosteric activator glucose 6-phosphate (G6P) was decreased in AMPKγ3(R200Q) relative to all other genotypes, whereas both AMPKγ3(R200Q) and AMPKγ3(R200Q) + RyR1(R615C) muscle exhibited increased GS activity with G6P. Increased activity of GS with G6P was not associated with increased abundance of GS or hexokinase 2. However, AMPKγ3(R200Q) enhanced UDP-glucose pyrophosphorylase 2 (UGP2) expression approximately threefold. Although UGP2 is not generally considered a rate-limiting enzyme for glycogen synthesis, our model suggests that UGP2 plays an important role in increasing flux to glycogen synthase. Moreover, we have shown that the capacity for glycogen storage is more closely related to the AMPKγ3(R200Q) mutation than activity.

Excess glycogen does not resolve high ultimate pH of oxidative muscle.

Skeletal muscle glycogen content can impact the extent of postmortem pH decline. Compared to glycolytic muscles, oxidative muscles contain lower glycogen levels antemortem which may contribute to the higher ultimate pH. In an effort to explore further the participation of glycogen in postmortem metabolism, we postulated that increasing the availability of glycogen would drive additional pH decline in oxidative muscles to equivalent pH values similar to the ultimate pH of glycolytic muscles. Glycolysis and pH declines were compared in porcine longissimus lumborum (glycolytic) and masseter (oxidative) muscles using an in vitro system in the presence of excess glycogen. The ultimate pH of the system containing longissimus lumborum reached a value similar to that observed in intact muscle. The pH decline of the system containing masseter samples stopped prematurely resulting in a higher ultimate pH which was similar to that of intact masseter muscle. To investigate further, we titrated powdered longissimus lumborum and masseter samples in the reaction buffer. As the percentage of glycolytic sample increased, the ultimate pH decreased. These data show that oxidative muscle produces meat with a high ultimate pH regardless of glycogen content and suggest that inherent muscle factors associated with glycolytic muscle control the extent of pH decline in pig muscles.

Net lactate accumulation and low buffering capacity explain low ultimate pH in the longissimus lumborum of AMPKγ3 R200Q mutant pigs.

Postmortem lactate accumulation in skeletal muscle is linearly associated with the extent of pH decline. Yet, pigs harboring the AMPKγ3(R200Q) mutation produce meat with similar lactate levels to that of wild-type pigs but have a lower ultimate pH. We hypothesized that lower initial lactate levels and (or) lower buffering capacity in muscle of these pigs may help explain this discrepancy. Longissimus lumborum muscle samples were harvested at 0 and 1440 min postmortem from AMPKγ3(R200Q) and wild-type pigs. As expected, AMPKγ3(R200Q) muscle exhibited a lower ultimate pH but similar lactate levels to that of wild-type pigs at 1440 min postmortem. However, the total net lactate produced postmortem was greater in the AMPKγ3(R200Q) muscle due to lower initial lactate levels at 0 min postmortem. Buffering capacity measured over the pH range of 5.5-7.0 was also lower in AMPKγ3(R200Q) muscle. Greater net lactate accumulation postmortem (i.e., glycolytic flux) coupled with a lower buffering capacity explains the lower ultimate pH of meat from AMPKγ3(R200Q) pigs.

Mitochondria influence postmortem metabolism and pH in an in vitro model.

Our objective was to determine the influence of mitochondria on metabolites and pH decline using an in vitro model of postmortem muscle metabolism. Mitochondria were isolated from porcine longissimus lumborum and added (0, 0.5, or 2.0mg) to powdered muscle in reaction media containing either a combination of inhibitors for mitochondria complexes (I, IV, and V) or diluent (without inhibitors). In the absence of inhibitors, adding mitochondria (0.5 and 2.0mg) reduced ATP loss from 30 to 120 min, but did not alter glycogen or lactate during this time. In reactions with mitochondria, inhibitors decreased ATP levels by 30 min and increased glycogen degradation by 60 min. Regardless of mitochondria content, inhibitors enhanced lactate accumulation from 15 to 240 min, and decreased pH from 15 min to 1440 min. In the in vitro model, mitochondria influence the maintenance of ATP, and inhibition of mitochondria enzyme activity contributes to accelerated metabolism and pH decline.

pH inactivation of phosphofructokinase arrests postmortem glycolysis.

Fresh meat quality development is influenced by pH decline that results from muscle glycolyzing energy substrates postmortem. The exact reason why glycolysis stops in the presence of residual glycogen remains unclear. We hypothesized that a critical glycolytic enzyme loses activity near the ultimate pH of meat. Porcine longissimus muscle samples were subjected to an in vitro system that mimics postmortem anaerobic metabolism at buffered pH values (7.0, 6.5, 6.0, 5.5 or 5.0). At pH7.0, 6.5, and 6.0, glycogenolysis and glycolysis proceeded normally while pH5.5 stopped lactate formation. Additional experimentation indicated that phosphofructokinase lost activity at pH5.5 while all other glycolytic enzymes remained active. A similar inactivation of phosphofructokinase was observed when using chicken and beef muscle. Elevated temperature hastened pH decline and phosphofructokinase activity loss. Thus, pH inactivates phosphofructokinase and arrests postmortem glycolysis, which may explain the similar ultimate pH across meat of different species.

Contribution of the phosphagen system to postmortem muscle metabolism in AMP-activated protein kinase γ3 R200Q pig Longissimus muscle.

Pigs with the AMP-activated protein kinase γ3 R200Q (AMPKγ3(R200Q)) mutation generate pork with low ultimate pH (pHu). We hypothesized that reducing muscle creatine (Cr) and phosphocreatine (PCr) may accelerate postmortem ATP consumption and prevent extended pH decline in AMPKγ3(R200Q) longissimus muscle. Wild type and AMPKγ3(R200Q) pigs were assigned to control diet or diet supplemented with the creatine analog ß-guanidinopropionic acid (ß-GPA, 1%) for 2 wk. ß-GPA reduced muscle PCr (P = 0.006) and total Cr (P<0.0001). In general, AMPKγ3(R200Q)+ß-GPA exhibited more rapid metabolism than control, AMPKγ3(R200Q), and ß-GPA treatment, evidenced by more rapid loss of ATP, more rapid increase in IMP, and decreased pH during the first 90 min postmortem. Overall, pHu was similar despite elevated glycogen (AMPKγ3(R200Q)), reduced total Cr (ß-GPA) or both (AMPKγ3(R200Q)+ß-GPA). Thus, reducing muscle phosphagens did not affect pHu in AMPKγ3(R200Q) muscle, but it hastened ATP depletion and pH decline.