4-hydroxy-3-methoxycinnamic acid (ferulic acid, FA) is known to have numerous beneficial health effects, including anti-obesity and anti-hyperglycemic properties. However, the molecular networks that modulate the beneficial FA-induced metabolic effects have not been well elucidated. In this study, we explored the molecular mechanisms mediating the beneficial metabolic effects of FA. In mice, FA protected against high-fat diet-induced weight gain, reduced food intake and exhibited an overall improved metabolic phenotype. The food intake suppression by FA was accompanied by a specific reduction in hypothalamic orexigenic neuropeptides, including agouti-related protein and neuropeptide Y, with no significant changes in the anorexigenic peptides pro-opiomelanocortin and cocaine and amphetamine-regulated transcript. FA treatment also inhibited fat accumulation in the liver and white adipose tissue and suppressed the expression of gluconeogenic genes, including phosphoenolpyruvate carboxylase and glucose-6-phosphatase. Furthermore, we show that FA phosphorylated and inactivated the transcription factor FoxO1, which positively regulates the expression of gluconeogenic and orexigenic genes, providing evidence that FA might exert its beneficial metabolic effects through inhibition of FoxO1 function in the periphery and the hypothalamus.

P2Y receptors are metabotropic G-protein-coupled receptors, which are involved in many important biologic functions in the central nervous system including retina. Subtypes of P2Y receptors in retinal tissue vary according to the species and the cell types. We examined the molecular and pharmacologic profiles of P2Y purinoceptors in retinoblastoma cell, which has not been identified yet. To achieve this goal, we used Ca2+ imaging technique and western blot analysis in WERI-Rb-1 cell, a human retinoblastoma cell line. ATP (10 microM) elicited strong but transient [Ca2+]i increase in a concentration-dependent manner from more than 80% of the WERI-Rb-1 cells (n=46). Orders of potency of P2Y agonists in evoking [Ca2+]i transients were 2MeS-ATP>ATP>>UTP=alphabeta-MeATP, which was compatible with the subclass of P2Y1 receptor. The [Ca2+]i transients evoked by applications of 2MeS-ATP and/or ATP were also profoundly suppressed in the presence of P2Y1 selective blocker (MRS 2179; 30 microM). P2Y1 receptor expression in WERI-Rb-1 cells was also identified by using western blot. Taken together, P2Y1 receptor is mainly expressed in a retinoblastoma cell, which elicits Ca2+ release from internal Ca2+ storage sites via the phospholipase C-mediated pathway. P2Y1 receptor activation in retinoblastoma cell could be a useful model to investigate the role of purinergic [Ca2+]i signaling in neural tissue as well as to find a novel therapeutic target to this lethal cancer.
Adenosine Triphosphate ; Blotting, Western ; Calcium ; Cell Line ; Central Nervous System ; Humans ; Phospholipases ; Receptors, G-Protein-Coupled ; Receptors, Purinergic P2Y ; Receptors, Purinergic P2Y1 ; Retina ; Retinaldehyde ; Retinoblastoma

In the heart, Na+-Ca2+ exchange (NCX) is the major Ca2+ extrusion mechanism. NCX has been considered as a relaxation mechanism, as it reduces global [Ca2+]i raised during activation. However, if NCX locates in the close proximity to the ryanodine receptor, then NCX would curtail Ca2+ before its diffusion to global Ca2+i. This will result in a global [Ca2+]i decrease especially during its ascending phase rather than descending phase. Therefore, NCX would decrease the myocardial contractility rather than inducing relaxation in the heart. This possibility was examined in this study by comparing NCX-induced extrusion of Ca2+ after its release from SR in the presence and absence of global Ca2+i transient in the isolated single rat ventricular myocytes by using patch-clamp technique in a whole-cell configuration. Global Ca2+i transient was controlled by an internal dialysis with different concentrations of BAPTA added in the pipette. During stimulation with a ramp pulse from +100 mV to -100 mV for 200 ms, global Ca2+i transient was suppressed only mildly, and completely at 1 mmol/L, and 10 mmol/L BAPTA, respectively. In these situations, ryanodine-sensitive inward NCX current was compared using 100micromol/L ryanodine, Na+ depletion, 5 mmol/L NiCl2 and 1micromol/L nifedipine. Surprisingly, the result showed that the ryanodine-sensitive inward NCX current was well preserved after 10 mmol/L BAPTA to 91 % of that obtained after 1 mmol/L BAPTA. From this result, it is concluded that most of the NCX-induced Ca2+ extrusion occurs before the Ca2+ diffuses to global Ca2+i in the rat ventricular myocyte.
Animals ; Architectural Accessibility ; Dialysis ; Diffusion* ; Heart ; Muscle Cells* ; Nifedipine ; Patch-Clamp Techniques ; Rats* ; Relaxation ; Ryanodine ; Ryanodine Receptor Calcium Release Channel

Voltage-sensitive release mechanism was pharmacologically dissected from the Ca2+-induced Ca2+ release in the SR Ca2+ release in the rat ventricular myocytes patch-clamped in a whole-cell mode. SR Ca2+ release process was monitored by using forward-mode Na+-Ca2+ exchange after restriction of the interactions between Ca2+ from SR and Na+-Ca2+ exchange within micro-domains with heavy cytosolic Ca2+ buffering with 10 mM BAPTA. During stimulation every 10 s with a pulse roughly mimicking action potential, the initial outward current gradually turned into a huge inward current of -12.9+/-0.5 pA/pF. From the inward current, two different inward INCXs were identified. One was 10 muM ryanodine-sensitive, constituting 14.2+/-2.3%. It was completely blocked by CdCl2 (0.1 mM and 0.5 mM) and by Na+-depletion. The other was identified by 5 mM NiCl2 after suppression of ICaL and ryanodine receptor, constituting 14.8+/-1.6%. This latter was blocked by either 10 mM caffeine-induced SR Ca2+-depletion or 1 mM tetracaine. IV-relationships illustrated that the latter was activated until the peak in 30~35 mV lower voltages than the former. Overall, it was concluded that the SR Ca2+ release process in the rat ventricular myocytes is mediated by the voltage-sensitive release mechanism in addition to the Ca2+-induced-Ca2+ release.
Action Potentials ; Animals ; Cadmium Chloride ; Cytosol ; Muscle Cells* ; Population Characteristics* ; Rats* ; Ryanodine Receptor Calcium Release Channel ; Tetracaine

Huge chondrosarcoma is a rare form of primary malignant tumor of the chest wall. We operated on a 60 year old female patient who had a huge anterior chest wall mass with local invasion into the pericardium and satellite tumors on the visceral pericardium of the heart. En-bloc resection of the huge tumor including both upper 3 ribs, both clavicles, manubrium of the sternum, pleura, and pericarium, was followed by complex chest wall reconstruction using a Gore-tex soft tissue patch and latissimus dorsi musculocutaneous free flap.
Chondrosarcoma* ; Clavicle ; Female ; Free Tissue Flaps ; Heart ; Humans ; Manubrium ; Mediastinal Neoplasms ; Middle Aged ; Pericardium ; Pleura ; Polytetrafluoroethylene ; Ribs ; Sternum ; Superficial Back Muscles ; Surgical Flaps ; Thoracic Wall* ; Thorax*

During depolarization, extrusion of Ca2+ from sarcoplasmic reticulum through forward-mode Na+ - Ca2+ exchange was studied in the rat ventricular myocytes patch-clamped in whole-cell configuration. In order to confine the Ca2+ responses in a micro-domain by limiting the Ca2+ diffusion time, rat ventricular myocytes were dialyzed with high (14 mM) EGTA. K+ current was suppressed by substituting KCl with 105 mM CsCl and 20 mM TEA in the pipette filling solution and by omitting KCl in the external Tyrode solution. Cl- current was suppressed by adding 0.1 mM DIDS in the external Tyrode solution. During stimulation roughly mimicking action potential, the initial outward current was converted into inward current, 47+/-1% of which was suppressed by 0.1 mM CdCl2. 10 mM caffeine increased the remaining inward current after CdCl2 in a cAMP-dependent manner. This caffeine-induced inward current was blocked by 1 muM ryanodine, 10 muM thapsigargin, 5 mM NiCl2, or by Na+ and Ca2+ omission, but not by 0.1 muM isoproterenol. The IapprxV relationship of the caffeine-induced current elicited inward current from -45 mV to +3 mV with the peak at -25 mV. Taken together, it is concluded that, during activation of the rat ventricular myocyte, forward-mode Na+ - Ca2+ exchange extrudes a fraction of Ca2+ released from sarcoplasmic reticulum mainly by voltage-sensitive release mechanism in a micro-domain in the t-tubule, which is functionally separable from global Cai2+ by EGTA.
4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid ; Action Potentials ; Animals ; Cadmium Chloride ; Caffeine ; Diffusion ; Egtazic Acid* ; Isoproterenol ; Muscle Cells* ; Rats* ; Ryanodine ; Sarcoplasmic Reticulum ; Tea ; Thapsigargin

Suppressive role of Na+-Ca2+ exchange in myocardial tension generation was examined in the negative frequency-force relationship (FFR) of electric field stimulated left atria (LA) from postnatal developing rat heart and in the whole-cell clamped adult rat ventricular myocytes with high concentration of intracellular Ca2+ buffer (14 mM EGTA). LA twitch amplitudes, which were suppressed by cyclopiazonic acid in a postnatal age-dependent manner, elicited frequency-dependent and postnatal age-dependent enhancements after Na+-reduced, Ca2+-depleted (26 Na-0 Ca) buffer application. These enhancements were blocked by caffeine pretreatment with postnatal age-dependent intensities. In the isolated rat ventricular myocytes, stimulation with the voltage protocol roughly mimicked action potential generated a large inward current which was partially blocked by nifedipine or Na+ current inhibition. 0 Ca application suppressed the inward current by 39 +/- 4% while the current was further suppressed after 0 Na-0 Ca application by 53 +/- 3%. Caffeine increased this inward current by 44 +/- 3% in spite of 14 mM EGTA. Finally, the Na+ current-dependent fraction of the inward current was increased in a stimulation frequency-dependent manner. From these results, it is concluded that the Ca2+ exit-mode (forward-mode) Na+-Ca2+ exchange suppresses the LA tension by extruding Ca2+ out of the cell right after its release from sarcoplasmic reticulum (SR) in a frequency-dependent manner during contraction, resulting in the negative frequency-force relationship in the rat LA.
Action Potentials ; Adult ; Animals ; Caffeine ; Calcium* ; Egtazic Acid ; Heart ; Humans ; Muscle Cells ; Myocardial Contraction* ; Nifedipine ; Rats* ; Sarcoplasmic Reticulum

The underlying mechanism commonly applicable for both the positive and negative force-frequency relationships (FFR) was pursued in left atria (LA) of rat and rabbit. The species differences in the roles of Na+/Ca2+ exchanger and sarcoplasmic reticulum (SR), which are major intracellular Ca2+ regulatory mechanisms in the heart, were examined in the amplitude accommodation to the frequency that changed from 3 Hz to the variable test frequencies for 5 minutes in the electrically field stimulated left atria (LA) of rat and rabbit. Norepinephrine strongly increased the frequency-related amplitude accommodation in both of rat and rabbit LA, while monensin, oubain or the reduced Na+ and 0 mM Ca2+ containing Tyrode solution increased the frequency-related amplitude accommodation only in the rabbit LA. Monenisn was also able to increase the frequency-related amplitude accommodation only in 1-day old rat LA but not in 4-week old rat LA that had 75% less Na+/Ca2+ exchanger with 97% higher SR than 1-day old rat LA. Taken together, it is concluded that the differences in the prevalence between myocardial Na+/Ca2+ exchanger and SR in the amplitude accommodation to the frequency-change determine the difference in the FFR between rat and rabbit heart.
Animals ; Heart ; Monensin ; Norepinephrine ; Prevalence ; Rats* ; Sarcoplasmic Reticulum

Frequency-force relationships (FFR) were studied in electrically field stimulated rat left atria (LA) by reducing the stimulation frequency from resting 3 Hz to test frequencies (0.1-1 Hz) for 5 minutes. The twitch amplitudes of LA elicited the typical negative staircases with 3-phased changes: the initial rapid increase, the second decrease and the following plateau at test frequencies. Verapamil (3 X 10-5 M) pretreatment elicited frequency-dependent suppression of the twitch amplitudes, exaggerating the negative staircase. Monensin pretreatment enhanced not the peak but the plateau amplitudes in a concentration-dependent manner. When the Na+-Ca2+ exchange was blocked by Na+ and Ca2+ depletion in the Krebs Hensleit buffer (0 Na+-0 Ca2+ KHB), the twitch amplitudes increased in a frequency-dependent manner, changing the negtive staircase into the positve one. Meanwhile, the 0 Na+-0 Ca2+ KHB applicationinduced enhancement was strongly suppressed by caffeine (5 mM) pretreatment. Only dibucaine among the local anesthetics increased the basal tone during frequency reduciton. There were no differences in 45Ca uptakes between 0.3 Hz and 3 Hz stimulation except at 1 min when it was significantly low at 0.3 Hz than 3 Hz, illustrating net Ca2+ losses. Monensin pretreatment enhanced the rate of this Ca2+ loss. Taken together, it is concluded that Na+-Ca2+ exchange extrudes more SR released Ca2+ out of the cell in proportion to the frequency, resulting in the negative rate staircase in the rat LA.
Anesthetics, Local ; Animals ; Caffeine ; Dibucaine ; Monensin ; Rats ; Verapamil

Recent reports revealed that the Na+/-Ca2+ exchangers and feet structures of sarcoplasmic reticulum (SR) are located in close vicinity in the specific compartment. Therefore, we investigated the possibility that the Na+/-Ca2+ exchanger may decrease the tension development by transporting the Ca2+ out of the cell right after it released from SR, on the basis of this anatomical proximity. We examined the negative force-frequency relationship of the developed tension in the electrically field stimulated left atria of postnatal developing rat (1, 3 day, 1 week and 4 week old after birth). Cyclopiazonic acid (3 X 10(-5) M) treatment decreased the developed tension further according to postnatal age. Monensin (3 X 10(-6) M) treatment did not increase the maximal tension in 4 week-old rat, preserving negative staircase, while the negative staircase in the younger rat were flattened. Ca2+ depletion in the buffer elicited more suppression of the maximal tension according to the frequency in all groups except the 4 week-old group. The % decrease of the maximal developed tension of 4 week-old group at 1 Hz to that of 0.1 Hz after Na+ and Ca2+ depletion was only a half of those of the younger groups. Taken together, it is concluded that the Na+/-Ca2+ exchange transports more Ca2+ released from SR out of the cell in proportion to the frequency, and this is responsible for the negative staircase effect of the rat heart.
Animals ; Foot ; Heart* ; Monensin ; Rats* ; Sarcoplasmic Reticulum