Involvement of multiple taste receptors in umami taste: analysis of gustatory nerve responses in metabotropic glutamate receptor 4 knockout mice.

KEY POINTS: The taste receptor T1R1 + T1R3 heterodimer and metabotropic glutamate receptors (mGluR) may function as umami taste receptors. Here, we used mGluR4 knockout (mGluR4-KO) mice and examined the function of mGluR4 in peripheral taste responses of mice. The mGluR4-KO mice showed reduced responses to glutamate and L-AP4 (mGluR4 agonist) in the chorda tympani and glossopharyngeal nerves without affecting responses to other taste stimuli. Residual glutamate responses in mGluR4-KO mice were suppressed by gurmarin (T1R3 blocker) and AIDA (group I mGluR antagonist). The present study not only provided functional evidence for the involvement of mGluR4 in umami taste responses, but also suggested contributions of T1R1 + T1R3 and mGluR1 receptors in glutamate responses. ABSTRACT: Umami taste is elicited by L-glutamate and some other amino acids and is thought to be initiated by G-protein-coupled receptors. Proposed umami receptors include heterodimers of taste receptor type 1, members 1 and 3 (T1R1 + T1R3), and metabotropic glutamate receptors 1 and 4 (mGluR1 and mGluR4). Accumulated evidences support the involvement of T1R1 + T1R3 in umami responses in mice. However, little is known about the in vivo function of mGluR in umami taste. Here, we examined taste responses of the chorda tympani (CT) and the glossopharyngeal (GL) nerves in wild-type mice and mice genetically lacking mGluR4 (mGluR4-KO). Our results indicated that compared to wild-type mice, mGluR4-KO mice showed significantly smaller gustatory nerve responses to glutamate and L-(+)-2-amino-4-phosphonobutyrate (an agonist for group III mGluR) in both the CT and GL nerves without affecting responses to other taste stimuli. Residual glutamate responses in mGluR4-KO mice were not affected by (RS)-alpha-cyclopropyl-4-phosphonophenylglycine (an antagonist for group III mGluR), but were suppressed by gurmarin (a T1R3 blocker) in the CT and (RS)-1-aminoindan-1,5-dicarboxylic acid (an antagonist for group I mGluR) in the CT and GL nerve. In wild-type mice, both quisqualic acid (an agonist for group I mGluR) and L-(+)-2-amino-4-phosphonobutyrate elicited gustatory nerve responses and these responses were suppressed by addition of (RS)-1-aminoindan-1,5-dicarboxylic acid and (RS)-alpha-cyclopropyl-4-phosphonophenylglycine, respectively. Collectively, the present study provided functional evidences for the involvement of mGluR4 in umami taste responses in mice. The results also suggest that T1R1 + T1R3 and mGluR1 are involved in umami taste responses in mice. Thus, umami taste would be mediated by multiple receptors.

Prevalence of anemia and iron deficiency and its association with body mass index in elite Japanese high school long-distance runners.

OBJECTIVES: Anemia is a common condition in long-distance runners (LDRs). Recently, not only iron deficiency (ID) but also energy deficiency has been considered as a risk factor for anemia in athletes but no evidence has yet been established. The aim of this study was to investigate the prevalence of anemia and ID and the influence of body mass index (BMI) on anemia in high-school LDRs. METHODS: The participants were 406 male and 235 female elite Japanese LDRs who competed in the All-Japan High-School Ekiden Championship 2019. They submitted their anthropometric data and results of a blood test within five days after the competition. The prevalence of anemia and ID and the influence of BMI on anemia were assessed retrospectively. RESULTS: Mean hemoglobin concentrations (Hb) were 14.8 ± 0.9 g/dl in males and 13.2 ± 0.9 g/dl in females. The prevalence of anemia (Hb < 14 g/dl in males and < 12 g/dl in females) was significantly higher in males (16.3%) than females (6.4%), but males also showed higher prevalence of non-iron deficiency anemia (NIDA) than females (11.6% and 3.0%, respectively). No significant gender difference was found in the prevalence of iron deficiency anemia (IDA) (4.7% in males and 3.4% in females). ID (serum ferritin level < 25 ng/ml) was significantly more prevalent in females (37.4%) than males (18.5%). A binary logistic regression analysis revealed that low BMI was a contributor to anemia in females (odds ratios: 0.577 (95% CI: 0.369-0.901), p = 0.012). CONCLUSION: In Japanese high-school LDRs, one in six males was anemic, but most males did not have ID. Conversely, one-third of females were diagnosed with ID. Lower BMI was identified as a risk for anemia in females, suggesting that leanness may also lead to anemia in females.

Use of nutritional supplements by elite Japanese track and field athletes.

BACKGROUND: While scientific evidence supports the efficacy of only limited nutritional supplements (NS) on sports performance, the use of NS is widespread in athletes. Given the serious issues of health damage or unintended Anti-Doping Rule Violations due to ingestion of contaminated NS in sports, accurately understanding NS practices by athletes is crucial. This study therefore elucidated the use of NS by elite Japanese track and field (TF) athletes. METHODS: The subjects were 574 Japanese TF athletes, including 275 junior athletes (under 20 years) and 299 senior athletes, who participated in international competitions from 2013 to 2018. Data on NS use were collected through pre-participation medical forms obtained from all entrants before their participation in competitions. NS users were requested to report the product names and primary components of all NS they were taking. RESULTS: The overall prevalence of NS use was 63.9%. The mean number of NS products used per athlete was 1.4. The prevalence was significantly higher in women (69.2%) than in men (59.6%) (p = 0.018) and significantly higher in senior athletes (68.9%) than in junior athletes (58.9%) (p = 0.012). The prevalence of NS use was higher in long-distance runners (75.8%) and lower in jumpers (52.3%) and throwers (49.2%) than other disciplines (p < 0.001). The most prevalent components were amino acids (49.3%), followed by vitamins (48.3%), minerals (22.8%), and protein (17.8%). CONCLUSIONS: Approximately two-thirds of elite Japanese TF athletes reported the use of NS, and NS practices varied by gender, age, and discipline.

A novel cardiac hypertrophic factor, neurotrophin-3, is paradoxically downregulated in cardiac hypertrophy.

The neurotrophin family plays pivotal roles in the development of the nervous system. Recently, the role of the neurotrophin in non-neural tissue has been extensively investigated. Among them, neurotrophin-3 and its receptor TrkC are critical for embryonic heart development, though little is known about neurotrophin-3/TrkC function in adult heart. Moreover, the expressions of other neurotrophin and Trk families in the cardiovascular system have not been fully determined. In adult and neonatal rats, only TrkC mRNA was expressed more abundantly in heart than aorta among the neurotrophin receptors, while all neurotrophins were equally expressed in the cardiovascular system. Immunohistochemistry confirmed the protein expressions of neurotrophin-3/TrkC in rat ventricles. In primary-cultured rat cardiomyocytes, neurotrophin-3 strongly activated p38 mitogen-activated protein kinase, extracellular signal-regulated kinase 1/2, and Jun N-terminal kinase pathways in Western blot analysis. In Northern blot analysis, neurotrophin-3 strongly increased mRNA expressions of cardiac hypertrophic markers (skeletal alpha-actin and atrial natriuretic peptide) in cardiomocytes. [(3)H]-phenylalanine uptake into cardiomyocytes, myofilament reorganization, and cardiomyocyte size were also augmented with neurotrophin-3 stimulation, indicating that neurotrophin-3 is a novel cardiac hypertrophic factor. Unexpectedly, neurotrophin-3 was downregulated in cardiac hypertrophy induced by pressure overload (in vivo), and in cardiomyocyte hypertrophy evoked by endothelin-1 stimulation (in vitro). Interestingly, the cell size and BNP mRNA expression level (markers of hypertrophy) were greater in cardiomyocytes treated with both neurotrophin-3 and endothelin-1 than in those stimulated with endothelin-1 alone. These findings demonstrate that neurotrophin-3 is a unique hypertrophic factor, which is paradoxically downregulated in cardiac hypertrophy and might counteract hypertrophic change.

The Vision and Challenges of Hokkaido Pharmaceutical University's Affiliated Pharmacy.

Hokkaido Pharmaceutical University (HPU), according to its educational mission, seeks to "develop medical professionals who contribute to community medicine", and it has produced more than 6300 graduates since 1974. With recent medical advancements and a progressively aging society, the role of the pharmacist in community medicine has diversified and is increasing in importance. Therefore, in April 2012, the Hokkaido Pharmaceutical University Affiliated Pharmacy was established as a for-profit business of the Educational Foundation of the Hokkaido University of Science, the parent body of HPU. The pharmacy is located near the Sapporo station; it is operated by six pharmacists and four clerks, and supported by three faculty members who are engaged in providing HPU student education such as on-site clinical training, in addition to their pharmacy duties such as home care pharmaceutics. For the first two years it was open, the pharmacy focused on the establishment of pharmacy administration and fiscal consolidation. In April 2015, the Pharmacy Management Committee set the pharmacy's future vision, as well as its mid-term strategy, which consists of the four main components of pharmacy practices, education, research, and social contribution, in order for the pharmacy to serve as a model of community pharmacy.

Bone marrow-derived regenerated cardiomyocytes (CMG Cells) express functional adrenergic and muscarinic receptors.

BACKGROUND: We recently reported that cardiomyocytes could be differentiated from bone marrow mesenchymal stem cells in vitro by 5-azacytidine treatment. In native cardiomyocytes, adrenergic and muscarinic receptors play crucial roles in mediating heart rate, conduction velocity, contractility, and cardiac hypertrophy. We investigated whether these receptors are expressed in differentiated CMG cells, and if so, whether they have downstream signaling systems. METHODS AND RESULTS: Reverse transcription-polymerase chain reaction revealed that CMG cells had already expressed alpha(1A)-, alpha(1B)-, and alpha(1D)-adrenergic receptor mRNA before 5-azacytidine treatment, whereas expression of beta(1)-, beta(2)-adrenergic and M(1)-, M(2)-muscarinic receptors was first detected at 1 day. Phenylephrine dose-dependently induced phosphorylation of ERK1/2, which was completely inhibited by prazosin, and significantly increased cell size. Isoproterenol augmented cAMP by 38-fold, which was fully inhibited by propranolol. Isoproterenol (10(-7) mol/L) increased the spontaneous beating rate by 47.6% (basal, 127+/-16 bpm), and propranolol and CGP20712A (beta(1)-selective blocker) reduced it by 79.0% and 71.0%, respectively, whereas ICI118551 (beta(2)-selective blocker) induced slight reduction. Cell motion, percent shortening, and contractile velocity were increased by 37.5%, 26.9%, and 50.6%, respectively, in response to isoproterenol. Phenylephrine and isoproterenol augmented ANP and BNP gene expressions. Carbachol increased IP(3) by 32-fold, which was markedly inhibited by atropine as well as AFDX116 (M(2)-selective blocker) measured by radioimmunoassay. CONCLUSIONS: These findings indicate that CMG cells expressed alpha(1A), alpha(1B), and alpha(1D) receptors before differentiation and expressed beta(1), beta(2), M(1), and M(2) receptors after they obtained the cardiomyocyte phenotype. These receptors had functional signal transduction pathways and could modulate cell function.

Marked improvement with sildenafil in a patient with primary pulmonary hypertension unresponsive to epoprostenol.

We report a 48-year-old woman with right heart failure due to primary pulmonary hypertension. Continuous infusion of epoprostenol (prostaglandin I2) for 1.5 years failed to control her condition, but she was later successfully treated with additional sildenafil for a few months. Her mean pulmonary artery pressure was originally 57 mmHg, increased to 62 mmHg with epoprostenol, and decreased to 45 mmHg with sildenafil. Additional sildenafil may be an effective and life-saving agent in patients with primary pulmonary hypertension who show a poor response to epoprostenol, which is considered to be very powerful medical treatment for the disease.

[Regenerative medicine in cardiology].

Emerging evidence indicates that cardiomyocytes can be regenerated from embryonic stem cells, adult mesenchymal stem cells, and cardiac side population cells (cardiac stem cells). When these cells were injected into diseased hearts, indices of cardiac function improved. There are ethical difficulties involved in the use of embryonic stem cells. The ratio of differentiation into cardiomyocytes is still relatively low at present. Transplantation of regenerated cardiomyocytes or stem cells, however, will become a future therapy for severe heart failure.

A case of transient mid-ventricular akinesia (a variant form of Takotsubo cardiomyopathy) followed with I-123-beta-metyl-iodophenyl pentadecanoic acid and I-123-meta-iodobenzyl-guanidine myocardial scintigraphy.

A 67-year-old woman without history of heart disease was admitted with chest oppression. Her electrocardiogram (ECG) at the time of admission showed ST segment elevation in leads V2-V6. Cardiac ultrasound revealed severe hypokinesis in mid to apical portion of anterior wall. Emergent coronary angiography showed normal coronary arteries. Left ventriculography (LVG) revealed akinesis of mid portion of anterior and inferior wall with hyperkinesis of apex and basal portion of anterior and inferior wall. Cardiac ultrasound examination 3 months later revealed improvement in LV contraction without mid-ventricular akisesia. The LVG performed 6 months later showed no focal asynergy. In I-123-beta-metyl-iodophenyl pentadecanoic acid myocardial scintigraphy the discrepancy of uptake between apical and anterior and inferior wall of mid region (more uptake in apex) was reduced. Using I-123-meta-iodobenzyl-guanidine myocardial scintigraphy in acute phase, decreased uptake in the mid portion of anterior and inferior to lateral wall was seen in early and delayed images and that persisted through 6 months. As these findings resembled those of Takotsubo cardiomyopathy other than affected region, it is possible to say that basically they belong to same entity of disease but they are different in their phenotype.

Oral sildenafil improves primary pulmonary hypertension refractory to epoprostenol.

BACKGROUND: Epoprostenol (prostaglandin I(2)) has become recognized as a therapeutic breakthrough that can improve hemodynamics and survival in patients with primary pulmonary hypertension (PPH). However, a significant number of patients have PPH that is refractory to epoprostenol, and lung transplantation has been the only remaining treatment option. METHODS AND RESULTS: The study subjects included 20 consecutive patients with PPH (mean pulmonary arterial pressure: 65+/-15 mmHg) who had received epoprostenol for more than 12 months. The patients were divided into 2 groups; responders and non-responders. In the non-responders, New York Heart Association (NYHA) functional class did not improve and mean right atrial pressure (mRA) increased to 8 mmHg or more, and additional sildenafil, a phosphodiesterase-5 inhibitor, was started. Six patients were included in the non-responders, whose mRA was 9+/-5 mmHg before and significantly increased to 13+/-3 mmHg after epoprostenol administration (p < 0.05). One patient died and the other 5 patients received oral sildenafil. The mRA of 12+/-4 mmHg (value before sildenafil) improved to 8+/-5 mmHg after sildenafil administration. Three patients were classified in the NYHA functional class 4 and improved to class 3, and 2 patients were in class 3 and remained in the same class after the addition of sildenafil. CONCLUSIONS: In patients with severe PPH refractory to epoprostenol treatment, additional oral sildenafil can improve pulmonary hemodynamics and symptoms. The combination therapy of epoprostenol and sildenafil is a new medical treatment to attempt before progressing to lung transplantation for patients with PPH refractory to epoprostenol.

Role of EGF Receptor and Pyk2 in endothelin-1-induced ERK activation in rat cardiomyocytes.

G protein-coupled receptor (GPCR)-evoked signal transduction pathways leading to the activation of extracellular signal-regulated kinases (ERK) are quite different among cell types. In cardiomyocytes, much attention has been focused on the activation of protein kinase C (PKC) or mobilization of intracellular Ca(2+) ([Ca(2+)](i)), however, the contributions of tyrosine kinases are controversial. In the present study, we characterized the signaling pathways involving tyrosine kinases, Pyk2 and epidermal growth factor receptor (EGFR), and their contribution to ERK activation in cultured cardiomyocytes. We initially investigated the potential involvement of [Ca(2+)](i) and PKC on the activation of these kinases in endothelin-stimulated cardiomyocytes. Interestingly, activation of Pyk2 was abrogated by chelating [Ca(2+)](i) or by downregulation of PKC, whereas transactivation of EGFR was solely dependent on PKC. By using a compound that selectively interferes with EGFR (AG1478), c-Src (PP1), or disrupts actin cytoskeleton (cytochalasin D), we demonstrated that cytochalasin D completely inhibited the activation of Pyk2, but not that of EGFR, whereas AG1478 did not inhibit the activation of Pyk2, indicating that transactivation of EGFR and signaling pathways involving Pyk2 were distinct pathways. Furthermore, activation of ERK and Shc, and c- fos gene expression were significantly inhibited by AG1478 but not by cytochalasin D or PP1. Overexpression of deletion mutant of EGFR attenuated the activation of ERK. These facts demonstrated the existence of two distinct tyrosine kinase pathways requiring Pyk2 or EGFR downstream from GPCR in cardiomyocytes. EGFR was Ca(2+)-independently activated and predominantly contributed to Shc/ERK/c- fos activation, while Pyk2 or c-Src contributed less to it.