ABSTRACT
The effects of phorbol-12,13-dibuterate (PDBu) on total sodium current (I(Na)-total), tetrodotoxin-resistant sodium current (I(Na)-TTXr), 4-AP-sensitive potassium current (I(A)) and TEA-sensitive potassium current (I(K)) in trigeminal ganglion (TG) neurons were investigated. Whole-cell patch clamp techniques were used to record ion currents in cultured TG neurons of rats. Results revealed that 0.5 micromol/L PDBu reduced the amplitude of I(Na)-total by (38.3+/-4.5)% (n=6, P0.05) nor the inactivation rate constant (control: 3.6+/-0.9 ms; PDBu: 3.6+/-0.8 ms; n=6, P>0.05) was altered. 0.5 micromol/L PDBu could significantly increase the amplitude of I(Na)-TTXr by (37.2+/-3.2)% (n=9, P0.05) or the inactivation rate constant (control: 4.6+/-0.6 ms; PDBu: 4.2+/-0.5 ms; n=5, P>0.05). 0.5 mumol/L PDBu inhibited I(K) by (15.6+/-5.0) % (n=16, P0.05). It was concluded that PDBu inhibited I(Na)-total but enhanced I(Na)-TTXr, and inhibited I(K) without affecting I(A). These data suggested that the activation of PKC pathway could exert the actions.
ABSTRACT
The effects of phorbol-12,13-dibuterate (PDBu) on total sodium current (INa-total), tetrodotoxin-resistant sodium current (INa-TTXr), 4-AP-sensitive potassium current (IA) and TEA-sensitive potassium current (IK) in trigeminal ganglion (TG) neurons were investigated.Whole-cell patch clamp techniques were used to record ion currents in cultured TG neurons of rats. Results revealed that 0.5 μmol/L PDBu reduced the amplitude of INa-total by (38.3±4.5)% (n=6, P＜0.05), but neither the G-V curve (control: V0.5 =-17.1±4.3 mV, k=7.4±1.3; PDBu: V0.5=-15.9±5.9 mV, k=5.9±1.4; n=6, P＞0.05) nor the inactivation rate constant (control: 3.6±0.9 ms; PDBu: 3.6±0.8 ms; n=6, P＞0.05) was altered. 0.5 μmol/L PDBu could significantly increase the amplitude of INa-TTXr by (37.2± 3.2)% (n=9, P＜0.05) without affecting the G-V curve (control: V0.5=-14.7±6.0 mV, k=6.9±1.4; PDBu: V0.5=-11.1±±5.3 mV, k=8.1±1.5; n=5, P＞0.05) or the inactivation rate constant (control: 4.6±±0.6 ms; PDBu: 4.2±0.5 ms; n=5, P＞0.05). 0.5 μmol/L PDBu inhibited IK by (15.6±5.0) % (n=16, P＜0.05), and V0.5 was significantly altered from - 4.7±1.4 mV to -7.9 ±1.8 mV (n=16, P＜0.05). IA was not significantly affected by PDBu, 0.5 μmol/L PDBu decreased IA by only (0.3±3.2)% (n=5, P＞0.05). It was concluded that PDBu inhibited INa-total but enhanced INa-TTXr, and inhibited IK without affecting IA. These data suggested that the activation of PKC pathway could exert the actions.
ABSTRACT
To further investigate the mechanisms of action of icariin (ICA), we assessed the effects of ICA on the in vitro formation of cGMP and cAMP in isolated rabbit corpus cavernosum. Isolated segments of rabbit corpus cavernosum were exposed to increasing concentrations of ICA and the dose-dependent accumulation of cGMP and cAMP was determined in the tissues samples by means of 125I radioimmunoassay. Responses of the isolated tissues preparations to ICA were compared with those obtained with the reference compounds sildenafil (Sild). Furthermore, the effects of ICA on the mRNA expression of specific cGMP-binding phosphodiesterase type V (PDE5) in rat penis were also observed. After incubation with ICA for 6 h or 14 h respectively, the levels of PDE5 mRNA were examined by reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that ICA increased cGMP concentrations directly (P 0.05). In the presence of sodium nitroprusside (SNP), a stimulatory agent of cGMP, both ICA and Sild increased cGMP concentrations with increasing dose (P 0.05). There were PDE5A1 and PDE5A2 mRNA expressions in rat corpus cavernosum with PDE5A2 being the dominant isoform. ICA could obviously inhibit these two isoforms mRNA expression in rat penis, and decrease PDE5A1 more pronouncedly (P < 0.01). The present study indicated that the aphrodisiac mechanisms of icariin involved the NO-cGMP signal transduction pathway, with increasing cGMP levels in the corpus cavernosum smooth muscle. The inhibitory effect of icariin on PDE5 mRNA expression, especially on PDE5A1, might account for its molecular mechanisms for its long-term activity.
ABSTRACT
To further investigate the mechanisms of action of icariin (ICA), we assessed the effects of ICA on the in vitro formation of cGMP and cAMP in isolated rabbit corpus cavernosum. Isolated segments of rabbit corpus cavernosum were exposed to increasing concentrations of ICA and the dose-dependent accumulation of cGMP and cAMP was determined in the tissues samples by means of 125I radioimmunoassay. Responses of the isolated tissues preparations to ICA were compared with those obtained with the reference compounds sildenafil (Sild). Furthermore, the effects of ICA on the mRNA expression of specific cGMP-binding phosphodiesterase type V (PDE5) in rat penis were also observed. After incubation with ICA for 6 h or 14 h respectively, the levels of PDE5 mRNA were examined by reverse transcriptase polymerase chain reaction (RT-PCR). The results showed that ICA increased cGMP concentrations directly (P＜0.05), but there was no significant effect on cAMP concentrations (P＞0.05). In the presence of sodium nitroprusside (SNP), a stimulatory agent of cGMP,both ICA and Sild increased cGMP concentrations with increasing dose (P＜0.01). Their EC50 was 4.62 (ICA) and 0.42 (Sild) μmol/L respectively. Under the same condition, ICA and Sild unaltered cAMP level significantly (P＞0.05). There were PDE5A1 and PDE5A2 mRNA expressions in rat corpus cavernosum with PDE5A2 being the dominant isoform. ICA could obviously inhibit these two isoforms mRNA expression in rat penis, and decrease PDE5A1 more pronouncedly (P＜ 0.01). The present study indicated that the aphrodisiac mechanisms of icariin involved the NO-cGMP signal transduction pathway, with increasing cGMP levels in the corpus cavernosum smooth muscle. The inhibitory effect of icariin on PDE5 mRNA expression, especially on PDE5A1, might account for its molecular mechanisms for its long-term activity.
ABSTRACT
Aim To explore the effects of lipoteichoic acid (LTA) induced delayed preconditioning (PC) on hypoxia-reoxygenation (H/R) injury of cultured human coronary artery endothelial cells (HCAECs), and to investigate the potential role of endogenous nitric oxide (NO) participated in the protective mechanism. Methods HCAECs were incubated for 2 h in a hypoxic atmosphere and reoxygenated for 4 h in a normoxic atmosphere. The delayed PC was induced by pretreatment with LTA assessed by the percentage of cellular injury with Trypan blue exclusion and by the amount of lactate dehydrogenase (LDH) in culture media. The NO level of the culture media was measured detect the expression of eNOS mRNA by RT-PCR method after cells were recovered from different points.Results LTA pretreatment significantly decreased the percentage of the killed cell and the concentration of LDH in media. Also, LTA pretreatment obviously raised the concentrations of NO in culture media. The protective effects of LTA were abrogated by pretreatment with N-monomethyl-L-arginine (L-NMMA).Moreover, the expression of eNOS mRNA was significantly upregulated after HCAECs exposure to LTA for 4 h following 2 h or 4 h recovery. Conclusion LTA could induce the delayed protection against H/R induced endothelial injury and dysfunction of cultured HCAECs. NO produced by eNOS acts initially as a trigger and subsequently as a mediator of delayed PC.
ABSTRACT
AIM To test if "adventitium-derived relaxing factor"(ADRF) possesses species- and tissue-specificity and make preliminary research on proteins separated from the bath solution. METHODS Record the tension of aortic ring with and without periadventitial fat, induced by phenylephrine(Phe) and analyze the proteins extracted from the bath solution with SDS-PAGE electrophoresis. RESULTS ① In Sprague-Dawley rats, the concentration-response curve of Phe to rings without the periadventitial fat shifted to rightward, as compared to the curve of the intact aortic rings, which means periadventitial fat can reduce the contraction induced by Phe. The same phenomena as the above could be found in aortic ring of Wistar rats, guinea pigs, and rabbits. ② Moreover, the contraction induced by Phe was obviously reduced by moving adipose tissue from greater omentum into the bath solution. ③ The release of ADRF was strongly reduced by 10 μmol·L-1 genistein (tyrosine kinase inhibitor). But the effect of existed ADRF could not be counterposed by genistein. ④ Five protein bands were separated from the bath solution, with relative molecular mass 74.0, 59.8, 54.4, 28.7 and 13.8 ku. CONCLUSION ① ADRF is a non-species specific factor. ② The entire name of ADRF should change from "adventitium-derived relaxing factor" to "adipocyte-derived relaxing factor". ③ Some proteins which may include ADRF are separated from the bath solution.
ABSTRACT
In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P > 0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal transduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.
Subject(s)
Berberine/pharmacology , Connective Tissue/physiopathology , Nitric Oxide Synthase/biosynthesis , Nitric Oxide Synthase/genetics , Nitric Oxide Synthase Type I/biosynthesis , Nitric Oxide Synthase Type I/genetics , Nitric Oxide Synthase Type III/biosynthesis , Nitric Oxide Synthase Type III/genetics , Penile Erection/physiology , Penis/metabolism , Penis/physiology , RNA, Messenger/biosynthesis , RNA, Messenger/geneticsABSTRACT
To investigate the effects of WIN 55,212-2 on IK in cultured rat trigeminal ganglion (TG) neurons, whole-cell patch clamp techniques were used to record the IK before and after WIN 55,212-2 perfusion at different concentrations. 30 μmol/L WIN 55,212-2 markedly (35.7 %±7.3%, P＜0. 01, n=8) inhibited IK currents, and the currents were partially recovered after washing.30μmol/L WIN 55,212-2 also induced a significant depolarizing shift in conductance-voltage parameters (control: V0.5 =10.43 ± 4.25 mV, k= 16.27±3.86; WIN 55,212-2: V0. 5 =24.71±3.91mV, k =16.69±2.75; n = 8, P＜0.01 for V0. 5). 0.01μmol/L WIN 55,212-2 slightly (27.0 %± 7.9 %, P＜0.05, n=7) increased IK currents, but had no significant change in conductance voltage parameters (control: V0.5 =10. 74±5. 27 mV, k=17. 33±2. 96; WIN 55,212-2: V0.5 =11.06±2.05 mV, k=19. 69±6. 60; n=7, P＞0.05 for V0.5 and k). These results suggested that WIN 55,212-2 has dual action, which might be through different receptors.
ABSTRACT
In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P＞0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal thansduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.
ABSTRACT
The hypoglycemic activity and its mechanism of Jatrorrhizine (Jat) were studied. The normal mice and alloxan-induced hyperglycemic mice were given with different doses of Jat. Blood glucose and liver glycogen levels were determined by spectrophotometry with glucose-oxidase and iodine reagents respectively. The levels of blood lactic acid (LC) and liver lactate dehydrogenase (LDH) activity were measured to explore the effect of Jat on anaerobic glycolysis. Succinate dehydrogenase (SDH) activity in liver was measured to evaluate the effect of Jat on aerobic glycolysis in liver. It was found that Jat (50 mg/kg, 100 mg/kg) could significantly decrease blood glucose level in a dose- and time-dependent manner in both normal and alloxan-diabetic mice, increase the activity of SDH, but had no significant effects on the LC level and LDH activity. Jat could significantly reduce the content of liver glycogen in normal mice. Moreover, Jat could inhibit the platelet aggregation in rabbits in vitro in a dose-effect relationship. It was concluded that Jat induced the pronounced decrease in blood glucose in normal and hyperglycemic mice. The hypoglycemic activity of Jat may be attributed to the enhancement of aerobic glycolysis.
ABSTRACT
<p><b>OBJECTIVE</b>To further investigate the action mechanisms of berberine (Ber) and to assess the effects of Ber on the mRNA expression of phosphodiesterase type 5 (PDE5) in rat corpus cavernosum.</p><p><b>METHODS</b>After incubating with Ber for 1 or 3 h respectively, we examined the levels of PDE5 mRNA by reverse transcription polymerase chain reaction (RT-PCR).</p><p><b>RESULTS</b>There were PDE5A1 and PDE5A2 mRNA expressions in the rat corpus cavernosum with PDE5A2 as the dominant isoform. Ber could obviously inhibit the mRNA expression of the two isoforms in the rat penis and bring on a pronounced decrease in PDE5A2 (P < 0.01).</p><p><b>CONCLUSION</b>The present study indicates that the inhibitory effect of Ber on PDE5 mRNA expression, especially on PDE5A2, might account for its molecular mechanism for treating ED.</p>
Subject(s)
Animals , Male , Rats , 3',5'-Cyclic-GMP Phosphodiesterases , Genetics , Berberine , Pharmacology , Cyclic Nucleotide Phosphodiesterases, Type 5 , Penis , Metabolism , RNA, Messenger , Rats, Wistar , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
To explore the potential of lipoteichoic acid (LTA) induced cardioprotection against ischemia-reperfusion (I/R) injury in isolated rat hearts and whether endogenous nitric oxide (NO) participates in the protection, the rats were pretreated with LTA (1 mg/kg, i.p.) 24 h before the experiment, and the isolated hearts were subjected to 30 min no-flow normothermic global ischemia and 60 min reperfusion after a 20-min stabilization period by the langendorff method. Cardiac functions were evaluated at the end of stabilization, and at 30 min, 60 min of reperfusion. The amounts of MB isoenzyme of creatine kinase (CK-MB), lactate dehydrogenase(LDH) and total NO oxidation products in the coronary effluent were measured spectrophotometrically at the end of reperfusion. It was revealed that pretreatment with LTA could significantly improve the recovery of cardiac function, reduce the release of CK-MB and LDH, and increase the concentrations of NO in coronary effluent. The protective effects were abrogated by pretreatment of the rats with L-NAME. It was concluded that LTA could induce the delayed cardioprotection against I/R injury, and endogenous NO may be involved in the mechanisms.
Subject(s)
Animals , Male , Rats , Cardiotonic Agents , Pharmacology , Creatine Kinase , Metabolism , Creatine Kinase, MB Form , In Vitro Techniques , Ischemic Preconditioning, Myocardial , Isoenzymes , Metabolism , L-Lactate Dehydrogenase , Metabolism , Lipopolysaccharides , Pharmacology , Myocardial Reperfusion Injury , Nitric Oxide , Metabolism , Rats, Wistar , Teichoic Acids , PharmacologyABSTRACT
The effects of Arecoline (Are) on calcium mobilization were investigated. In isolated single ventricular myocyte of guinea pig, patch clamp whole cell recording techniques were used to record the current of L-type calcium channel and cytosolic Ca2+ level ([Ca2+]i) labeled with fluorescence probe Fluo-3/AM was measured under a laser scanning confocal microscope. Results revealed that Are (3-100 mumol/L) could inhibit L-type calcium current in a concentration-dependent manner and the value of IC50 was 33.73 mumol/L (n = 5). In the absence of extracellular calcium, the resting levels of [Ca2+]i was not affected by Are (n = 6, P > 0.05), but pretreatment with Are (30 mumol/L) could significantly inhibit the [Ca2+]i elevation induced by caffeine (10 mmol/L, n = 6, P < 0.01). It was concluded that Are could inhibit not only calcium influx through L-type calcium channel but also calcium release from sarcoplasmic reticulum.