Secretion of one adipokine Nampt/Visfatin suppresses the inflammatory stress-induced NF-κB activity and affects Nampt-dependent cell viability in Huh-7 cells.

Nampt/visfatin acts in both intracellular and extracellular compartments to regulate multiple biological roles, including NAD metabolism, cancer, inflammation, and senescence. However, its function in chronic inflammation and carcinogenesis in hepatocellular carcinoma (HCC) has not been well-defined. Here we use Huh-7 hepatoma cells as a model to determine how Nampt/visfatin affects cellular survival under oxidative stress. We found that the transition of Nampt/visfatin from intracellular into extracellular form was induced by H2O2 treatment in 293T cells and confirmed that this phenomenon was not due to cell death but through the secretion of Nampt/visfatin. In addition, Nampt/visfatin suppressed cell viability in oxidative treatment in Huh-7 cells and acted on the inhibition of hepatoma cell growth. Oxidative stress also reduced the Nampt-mediated activation of NF-κB gene expression. In this study, we identify a novel feature of Nampt/visfatin which functions as an adipokine that can be secreted upon cellular stress. Our results provide an example to understand how adipokine interacts with chemotherapeutic treatment by oxidative stress in HCC.

Identification of nucleotide sequence involved in Weissellicin L production.

BACKGROUND: Weissellicin L, a novel bacteriocin produced by Weissella hellenica 4-7, was previously characterized but its full amino acid sequence remain unknown. The draft genome sequencing analysis of Weissella hellenica 4-7 was performed and the open reading frame (ORF) encoding the weissellicin L was identified and clarified. FINDINGS: The obtained results indicated that the mature bacteriocin consists of 29 amino acid residues with a molecular weight of approximately 3205.64 Da. A conserved processing site of two glycine residues in positions -1 and -2 was observed in the leader peptides. The possibility that bacteriocin secretion depended on ATP-binding cassette (ABC) transporter was therefore suggested. Furthermore, primers were designed from 5' and 3' flanking sequences of the weissellicin L structural gene. PCR presented a single product and was useful to detect weissellicin L structural gene. CONCLUSIONS: To our knowledge, this is the first report describing the full amino acid sequence of Weissellicin L. A rapid method to detect weissellicin L structural gene was also reported in this study.

Inhibition of peroxisome proliferator-activated receptor gamma prevents the melanogenesis in murine B16/F10 melanoma cells.

The purpose of this study was to investigate if PPARγ plays a role in the melanogenesis. B16/F10 cells were divided into five groups: control, melanin stimulating hormone (α-MSH), α-MSH+retinol, α-MSH+GW9662 (PPARγ antagonist), and GW9662. Cells in the control group were cultured in the Dulbecco's modified Eagle's medium (DMEM) for 48 hrs. To initiate the melanogenesis, cells in all α-MSH groups were cultured in medium containing α-MSH (10 nM) for 48 hrs. Cells were treated simultaneously with retinol (5 µM) in the α-MSH+retinol group. Instead of retinol, GW9662 (10 µM) was cocultured in the α-MSH+GW9662 group. Cells in the final group were cultured in the DMEM with GW9662. All the analyses were carried out 48 hours after treatments. The α-MSH was able to increase cell number, melanin production, and the activity of tyrosinase, the limiting enzyme in melanogenesis. These α-MSH-induced changes were prevented either by retinol or by GW9662. Further analyses of the activities of antioxidant enzymes including glutathione, catalase, and the superoxide dismutase (SOD) showed that α-MSH treatment raised the activity of SOD which was dependent on PPARγ level. According to our results, the α-MSH-induced melanogenesis was PPARγ dependent, which also modulated the expression of SOD.

Lactococcus formosensis sp. nov., a lactic acid bacterium isolated from yan-tsai-shin (fermented broccoli stems).

A coccal-shaped organism, designated 516(T), was isolated from yan-tsai-shin (fermented broccoli stems), a traditional fermented food in Taiwan. 16S rRNA gene sequencing results showed that strain 516(T) had 98.9 % sequence similarity to that of the type strain Lactococcus garvieae NBRC 100934(T). Comparison of three housekeeping genes, rpoA, rpoB and pheS, revealed that strain 516(T) was well separated from Lactococcus garvieae NBRC 100934(T). DNA-DNA hybridization studies indicated that strain 516(T) had low DNA relatedness with Lactococcus garvieae NBRC 100934(T) (46.1 %). The DNA G+C content of strain 516(T) was 38.1 mol% and the major fatty acids were C16 : 0 (22.7 %), C19 : 0 cyclo ω8c (17.9 %) and summed feature 7 (29.0 %). Based on the evidence, strain 516(T) represents a novel species of the genus Lactococcus, for which the name Lactococcus formosensis sp. nov. is proposed. The type strain is 516(T) ( = NBRC 109475(T) = BCRC 80576(T)).

Diversity of lactic acid bacteria associated with fresh coffee cherries in Taiwan.

A total of 102 lactic acid bacteria (LAB) were isolated from three different coffee farms in Taiwan. These isolates were classified and identified by the restriction fragment length polymorphism analysis and sequencing of 16S ribosomal DNA. Heterofermentative Leuconostoc, and Weissella species were the most common LAB found in two farms located at an approximate altitude of 800 m. Lactococcus lactis subsp. lactis was the most common LAB found in the remaining farm was located at an approximate altitude of 1,200 m. It is therefore suggested that the altitude and climate may affect the distribution of LAB. On the basis of phylogenetic analysis, two strains included in the genera Enterococcus were considered as two potential novel species or subspecies. In addition, a total of 34 isolates showed the antifungal activity against Aspergillus flavus. Moreover, seven Lactococcus lactis subsp. lactis strains and one Enterococcus faecalis strain were found to have bacteriocin-like inhibitory substance-producing capability. These results suggest that various LAB are associated with fresh coffee cherries in Taiwan. Some of the isolates found in this study showed potential as antifungal agents.

Enterocin T, a novel class IIa bacteriocin produced by Enterococcus sp. 812.

Enterococcus sp. 812, isolated from fresh broccoli, was previously found to produce a bacteriocin active against a number of Gram-positive bacteria, including Listeria monocytogenes. Bacteriocin activity decreased slightly after autoclaving (121 °C for 15 min), but was inactivated by protease K. Mass spectrometry analysis revealed the bacteriocin mass to be approximately 4,521.34 Da. N-terminal amino acid sequencing yielded a partial sequence, NH2-ATYYGNGVYXDKKKXWVEWGQA, by Edman degradation, which contained the consensus class IIa bacteriocin motif YGNGV in the N-terminal region. The obtained partial sequence showed high homology with some enterococcal bacteriocins; however, no identical peptide or protein was found. This peptide was therefore considered to be a novel bacteriocin produced by Enterococcus sp. 812 and was termed enterocin T.

Enterococcus saccharolyticus subsp. taiwanensis subsp. nov., isolated from broccoli.

A coccal strain isolated from fresh broccoli was initially identified as Enterococcus saccharolyticus; however, molecular identification and phenotypic traits did not support this identification. DNA-DNA hybridization with the type strain of E. saccharolyticus (76.4 % relatedness), DNA G+C content (35.7 mol%), phylogenetic analysis based on 16S rRNA, pheS and rpoA gene sequences, rep-PCR fingerprinting and profiles of cellular fatty acids, whole-cell proteins and enzyme activities, together with carbohydrate metabolism characteristics, indicated that this strain is distinct and represents a novel subspecies, for which the name Enterococcus saccharolyticus subsp. taiwanensis subsp. nov. is proposed. The type strain is 812(T) ( = NBRC 109476(T) = BCRC 80575(T)). Furthermore, we present an emended description of Enterococcus saccharolyticus and proposal of Enterococcus saccharolyticus subsp. saccharolyticus subsp. nov. (type strain ATCC 43076(T) = CCUG 27643(T) = CCUG 33311(T) = CIP 103246(T) = DSM 20726(T) = JCM 8734(T) = LMG 11427(T) = NBRC 100493(T) = NCIMB 702594(T)).

Draft Genome Sequence of Lactobacillus pobuzihii E100301T.

Lactobacillus pobuzihii E100301(T) is a novel Lactobacillus species previously isolated from pobuzihi (fermented cummingcordia) in Taiwan. Phylogenetically, this strain is closest to Lactobacillus acidipiscis, but its phenotypic characteristics can be clearly distinguished from those of L. acidipiscis. We present the draft genome sequence of strain L. pobuzihii E100301(T).

Lactococcus taiwanensis sp. nov., a lactic acid bacterium isolated from fresh cummingcordia.

One coccal strain, designated 0905C15(T), was isolated from fresh cummingcordia, which is the main ingredient of pobuzihi (fermented cummingcordia), a traditional fermented food in Taiwan. 16S rRNA gene sequencing results showed that strain 0905C15(T) had 98.22-98.82 % sequence similarity to that of the type strains of four Lactococcus lactis subspecies (L. lactis subsp. lactis BCRC 12312(T), L. lactis subsp. cremoris BCRC 12586(T), L. lactis subsp. hordniae BCRC 80474(T) and L. lactis subsp. tructae BCRC 80475(T)). Comparison of two housekeeping genes, recA and rpoB, revealed that strain 0905C15(T) was well separated from the reference strains of the genus Lactococcus. DNA-DNA hybridization studies indicated that strain 0905C15(T) had low DNA relatedness to the four Lactococcus lactis subspecies (9.7-15.24 %). The DNA G+C content of strain 0905C15(T) was 39.6 mol %. Based on the evidence, strain 0905C15(T) represents a novel species of the genus Lactococcus, for which the name Lactococcus taiwanensis sp. nov. is proposed. The type strain is 0905C15(T) ( = NBRC 109049(T) = BCRC 80460(T)).

Attenuation of the extract from Moringa oleifera on monocrotaline-induced pulmonary hypertension in rats.

The purpose of this study was to determine the effects of an extract from Moringa oleifera (MO) on the development of monocrotaline (MCT)-induced pulmonary hypertension (PH) in Wistar rats. An ethanol extraction was performed on dried MO leaves, and HPLC analysis identified niaziridin and niazirin in the extract. PH was induced with a single subcutaneous injection of MCT (60 mg/kg) which resulted in increases in pulmonary arterial blood pressure (Ppa) and in thickening of the pulmonary arterial medial layer in the rats. Three weeks after induction, acute administration of the MO extract to the rats decreased Ppa in a dose-dependent manner that reached statistical significance at a dose of 4.5 mg of freeze-dried extract per kg body weight. The reduction in Ppa suggested that the extract directly relaxed the pulmonary arteries. To assay the effects of chronic administration of the MO extract on PH, control, MCT and MCT+MO groups were designated. Rats in the control group received a saline injection; the MCT and MCT+MO groups received MCT to induce PH. During the third week after MCT treatment, the MCT+MO group received daily i.p. injections of the MO extract (4.5 mg of freeze-dried extract/kg of body weight). Compared to the control group, the MCT group had higher Ppa and thicker medial layers in the pulmonary arteries. Chronic treatments with the MO extract reversed the MCT-induced changes. Additionally, the MCT group had a significant elevation in superoxide dismutase activity when normalized by the MO extract treatments. In conclusion, the MO extract successfully attenuated the development of PH via direct vasodilatation and a potential increase in antioxidant activity.

Acute neurosteroids inhibit the spinal reflex potentiation via GABAergic neurotransmission.

Recently, we demonstrated a chronic neurosteroid-dependent inhibition of activity-dependent spinal reflex potentiation (SRP), but it remains unclear whether neurosteroids acutely modulate SRP induction. This study shows progesterone as well as two of its 3alpha,5alpha-derivatives, allopregnalonone and 3alpha,5alpha-tetrahydrodeoxycorticosterone (THDOC), to be capable of producing acute GABA(A) receptor (GABA(A)R)-dependent inhibition of SRP. When compared with test simulation (1 stimulation/30 s) of pelvic afferent nerves that evoked a baseline reflex activity in an external urethra sphincter electromyogram, repetitive stimulation (RS; 1 stimulation/1 s) induced SRP characterized by an increase in the evoked activity. Intrathecal progesterone (3-30 muM, 10 microl) at 10 min before stimulation onset dose dependently prevented RS induction. Intrathecal allopregnalonone (10 muM, 10 microl it) and THDOC (10 microM, 10 microl it) also prevented the SRP caused by RS. Pretreatment with the GABA(A)R antagonist bicuculline (10 microM, 10 microl it) at 1 min before progesterone/neurosteroid injection attenuated the inhibition of SRP caused by progesterone, allopregnanolone, and THDOC. Results suggest that progesterone and its neurosteroid metabolites may be crucial to the development of pelvic visceral neuropathic/postinflammatory pain and imply clinical use of neurosteroids, such as allopregnanolone and THDOC, for visceral pain treatment.

Nicotine-activated descending facilitation on spinal NMDA-dependent reflex potentiation from pontine tegmentum in rats.

This study was conducted to investigate the possible neurotransmitter that activates the descending pathways coming from the dorsolateral pontine tegmentum (DPT) to modulate spinal pelvic-urethra reflex potentiation. External urethra sphincter electromyogram (EUSE) activity in response to test stimulation (TS, 1/30 Hz) and repetitive stimulation (RS, 1 Hz) on the pelvic afferent nerve of 63 anesthetized rats were recorded with or without microinjection of nicotinic cholinergic receptor (nAChR) agonists, ACh and nicotine, to the DPT. TS evoked a baseline reflex activity with a single action potential (1.00 +/- 0.00 spikes/stimulation, n = 40), whereas RS produced a long-lasting reflex potentiation (16.14 +/- 0.96 spikes/stimulation, n = 40) that was abolished by d-2-amino-5-phosphonovaleric acid (1.60 +/- 0.89 spikes/stimulation, n = 40) and was attenuated by 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F) quinoxaline (7.10 +/- 0.84 spikes/stimulation, n = 40). ACh and nicotine microinjections to DPT both produced facilitation on the RS-induced reflex potentiation (23.57 +/- 2.23 and 28.29 +/- 2.36 spikes/stimulation, P < 0.01, n = 10 and 20, respectively). Pretreatment of selective nicotinic receptor antagonist, chlorisondamine, reversed the facilitation on RS-induced reflex potentiation caused by nicotine (19.41 +/- 1.21 spikes/stimulation, P < 0.01, n = 10) Intrathecal WAY-100635 and spinal transection at the T(1) level both abolished the facilitation on reflex potentiation resulting from the DPT nicotine injection (12.86 +/- 3.13 and 15.57 +/- 1.72 spikes/stimulation, P < 0.01, n = 10 each). Our findings suggest that activation of nAChR at DPT may modulate N-methyl-d-aspartic acid-dependent reflex potentiation via descending serotonergic neurotransmission. This descending modulation may have physiological/pathological relevance in the neural controls of urethral closure.

Calcium/calmodulin-dependent kinase II mediates NO-elicited PKG activation to participate in spinal reflex potentiation in anesthetized rats.

Calcium/calmodulin protein kinase (CaMK)-dependent nitric oxide (NO) and the downstream intracellular messenger cGMP, which is activated by soluble guanylate cyclase (sGC), are believed to induce long-term changes in efficacy of synapses through the activation of protein kinase G (PKG). The aim of this study was to examine the involvement of the CaMKII-dependent NO/sGC/PKG pathway in a novel form of repetitive stimulation-induced spinal reflex potentiation (SRP). A single-pulse test stimulation (TS; 1/30 Hz) on the afferent nerve evoked a single action potential, while repetitive stimulation (RS; 1 Hz) induced a long-lasting SRP that was abolished by a selective Ca(2+)/CaMKII inhibitor, autocamtide 2-related inhibitory peptide (AIP). Such an inhibitory effect was reversed by a relative excess of nitric oxide synthase (NOS) substrate, L-arginine. In addition, the RS-induced SRP was abolished by pretreatment with the NOS inhibitor, N(G)-nitro-L-arginine-methyl ester (L-NAME). The sGC activator, protoporphyrin IX (PPIX), reversed the blocking effect caused by L-NAME. On the other hand, a sGC blocker, 1H-[1, 2, 4]oxadiazolo[4, 3-alpha]quinoxalin-1-one (ODQ), abolished the RS-induced SRP. Intrathecal applications of the membrane-permeable cGMP analog, 8-bromoguanosine 3',5'-cyclic monophosphate sodium salt monohydrate (8-Br-cGMP), reversed the blocking effect on the RS-induced SRP elicited by the ODQ. Our findings suggest that a CaMKII-dependent NO/sGC/PKG pathway is involved in the RS-induced SRP, which has pathological relevance to hyperalgesia and allodynia.

Descending facilitation of spinal NMDA-dependent reflex potentiation from pontine tegmentum in rats.

This study was conducted to investigate whether dorsolateral pontine tegmentum stimulation modulates spinal reflex potentiation (SRP) and whether serotonergic neurotransmission is involved in such a modulation. Reflex activities of the external urethra sphincter (EUS) electromyogram in response to a test stimulation (TS; 1/30 Hz) or repetitive stimulation (RS; 1 Hz) on the pelvic afferent nerve in 35 anesthetized rats were recorded with/without synchronized train pontine stimulation (PS; 300 Hz, 30 ms) and/or intrathecal administrations of 10 microl of 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F) quinoxaline (NBQX; 100 microM), d-2-amino-5-phosphonovalerate (APV; 100 microM), N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexanecarboxamide trihydrochloride (WAY 100635; 100 microM), and 8-hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT; 100 microM). The TS evoked a single action potential (1.00 +/- 0.00 spikes/stimulation), while the RS produced a long-lasting SRP (16.12 +/- 1.59 spikes/stimulation) that was abolished by APV (1.57 +/- 0.29 spikes/stimulation) and was attenuated by NBQX (7.42 +/- 0.57 spikes/stimulation). Synchronized train PS with RS (PS+RS) produced facilitation in RS-induced SRP (25.17 +/- 2.21 spikes/stimulation). Intrathecal WAY 100635 abolished the facilitation in SRP as a result of the synchronized PS (14.66 +/- 1.58 spikes/stimulation). On the other hand, intrathecal 8-OH-DPAT elicited facilitation in the RS-induced SRP (25.16 +/- 1.05 spikes/stimulation) without synchronized PS. Our findings suggest that dorsolateral pontine tegmentum may modulate N-methyl-d-aspartic acid-dependent SRP via descending serotonergic neurotransmission. This descending modulation may have physiological/pharmacological relevance in the neural controls of urethral closure.

Spinal glutamatergic NMDA-dependent cyclic pelvic nerve-to-external urethra sphincter reflex potentiation in anesthetized rats.

The purposes of this study were to investigate whether the pelvic nerve-to-external urethra sphincter (EUS) reflex potentiation can be induced under physiological conditions and to determine whether glutamatergic neurotransmission is involved in the reflex potentiation. Stimulation-evoked reflex activities, during rhythmic bladder contractions caused by a continuous saline infusion, in 21 anesthetized rats were recorded with/without the intrathecal administration of 10 microl of CNQX (a glutamatergic AMPA receptor antagonist; 100 microM) and APV( a glutamatergic NMDA receptor antagonist; 100 microM). Reflex activities became potentiated following the increment of intravesical pressure (IVP) during the storage phase (2.39 +/- 0.28 spikes/mmHg, n = 21) and the ascending period of the voiding phase (1.46 +/- 0.35 spikes/mmHg, n = 21) and decreased following the decrement of IVP during the descending period of the voiding phase (1.50 +/- 0.33 spikes/mmHg, n = 21). Although it is characterized by a low IVP, a postvoiding reflex potentiation in stimulation-evoked activities was elicited at the critical period after a voiding contraction had just finished (23.95 +/- 8.96 spikes/mmHg, n = 21). The slope of the regression line of evoked activities vs. the IVP during the storage phase was significantly (P < 0.01) higher than that of the ascending and descending periods of the voiding phase, but there was no statistical difference between the ascending and the descending periods (P > 0.05). In addition, the slope of the regression line of posttetanic reflex potentiation was significantly higher than that of the storage phase (P < 0.01). All the slopes of the regression lines decreased after intrathecal CNQX administration (from 3.15 +/- 0.44, 2.10 +/- 0.57, 2.13 +/- 0.53, and 21.30 +/- 3.41 to 0.83 +/- 0.31, 0.74 +/- 0.12, 0.76 +/- 0.12, and 4.31 +/- 3.71 spikes/mmHg in storage, ascending and descending period of the voiding phase, and postvoiding potentiation, respectively; all P < 0.01, n = 10). The slopes of the regression lines became almost horizontal after intrathecal APV administration (from 3.15 +/- 0.44, 2.10 +/- 0.57, 2.13 +/- 0.53, and 21.30 +/- 3.41 to 0.16 +/- 0.12, 0.21 +/- 0.07, 0.18 +/- 0.05, and 0.23 +/- 0.76 spikes/mmHg in storage, ascending and descending period of voiding phase, and postvoiding potentiation, respectively; all P < 0.01, n = 10). Our results suggest that a potentiation in the pelvic nerve-to-EUS reflex can be induced under physiological conditions and the glutamatergic mechanism appears to be involved in this reflex potentiation.

Spinal glutamatergic NMDA-dependent pelvic nerve-to-external urethra sphincter reflex potentiation caused by a mechanical stimulation in anesthetized rats.

The current study investigates whether the spinal pelvic nerve-to-external urethra sphincter (EUS) reflex potentiation can be induced by a mechanical stimulation and whether the glutamatergic mechanism is involved in yielding such a reflex potentiation. The external urethra sphincter electromyogram (EUSE) activity, evoked by a single or by repetitive pelvic nerve stimulation, in 30 anesthetized rats was recorded with/without bladder saline distension. Without saline distension (0 cmH(2)O), a single pulse nerve stimulation evoked a single action potential in the reflex activity, whereas repetitive pelvic stimulation and saline distension (6 approximately 20 cmH(2)O) both elicited a long-lasting reflex potentiation (20.05 +/- 3.21 and 75.01 +/- 9.87 spikes/stimulation, respectively). The saline distension-induced pelvic nerve-to-EUS reflex potentiation was abolished by D-2-amino-5-phosphonovalerate [APV; a glutamatergic N -methyl-D-aspartic acid (NMDA) receptor antagonist; 100 microM, 10 microl, 1.72 +/- 0.31 spikes/stimulation] and attenuated by 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F) quinoxaline [NBQX; a glutamatergic alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionate (AMPA) receptor antagonist; 100 microM, 10 microl, 26.16 +/- 7.27 spikes/stimulation], but was not affected by bicuculline (a GABAergic antagonist; 100 microM, 10 microl, 53.62 +/- 15.54 spikes/stimulation). Intrathecal administration of glutamate (31.12 +/- 8.25 spikes/stimulation, 100 microM, 10 microl) and NMDA (26.25 +/- 4.12 spikes/stimulation, 100 microM, 10 microl) both induced a long-lasting pelvic nerve-to-EUS reflex potentiation without saline distension, which was similar to the findings observed from saline distension only. The duration of the contraction wave of the urethra was elongated by the saline distension-induced pelvic nerve-to-EUS reflex potentiation, whereas the peak pressure of the contraction wave was not affected. Our findings suggest that saline distension in the bladder elicits a pelvic nerve-to-EUS reflex potentiation and the glutamatergic mechanism contributes to the presence of such a reflex potentiation.

Effects of selective unilateral dorsal root(s) rhizotomy on micturition reflex in anesthetized rats.

PURPOSE: To clarify the contributions of sensory inputs and glutamate transmissions to the spinal micturition reflex. MATERIALS AND METHODS: Cystometrogram and external urethral sphincter electromyogram activities evaluated the L6 and/or S1 levels. RESULTS: Changes in intravesicular pressure (IVP) in response to saline infusion (0.1 ml/min) were found after unilateral dorsal root rhizotomy at the L6 level, which showed significant increases in threshold pressure (rhizotomized vs. control: 14.25 +/- 0.82 vs. 8.40 +/- 0.69 cmH(2)O, P < 0.01, n = 28), post-voiding pressure (7.66 +/- 0.56 vs. 5.42 +/- 0.52 cmH(2)O, P < 0.01, n = 28), holding duration (135.06 +/- 23.6 vs. 77.73 +/- 13.56 sec, P < 0.05, n = 28), and inter-contraction interval (140.62 +/- 23.29 vs. 82.40 +/- 13.57 sec, P < 0.05, n = 28). Several (mean = 2.32 +/- 1.31 vs. 0.12 +/- 0.21, P < 0.01, n = 28, P < 0.01, n = 28) non-voiding contractions with gradual increase in IVP were found ahead of voiding contraction after rhizotomy. An additional dorsal root rhizotomy at the ipsilateral S1 level caused further increases in urodynamic parameters (threshold pressure, 18.18 +/- 1.67 cmH(2)O, P < 0.01; post-voiding pressure 8.07 +/- 0.96 cmH(2)O, P < 0.01; holding duration, 211.44 +/- 42.54 sec, P < 0.01; inter-contraction interval, 264.2 +/- 59.99 sec, P < 0.05; non-voiding contractions, 4.41 +/- 2.12, P < 0.01, n = 7). Intrathecal glutamate (100 microM, 10 microl) ameliorated all the pathological conditions induced by unilateral dorsal root rhizotomy at the L6 level in a dose dependent manner (ED(50) = 1.25 +/- 10(-5)). Intrathecal CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; 100 microM, 10 microl) and APV (D-2-amino-5-phosphonovaleric acid; 100 microM, 10 microl) injections after rhizotomy at the L6 level induced disturbances similar to that caused by an additional rhizotomy at ipsilateral S1 level. Wherease, glutamate (100 microM, 10 microl) reversed the disturbances caused by CNQX but showed no effect on that by APV. CONCLUSIONS: Acute partial sensory deprivation caused acute impaired micturition reflex in rat models. In addition, glutamatergic NMDA and AMPA receptors are important for mediating these impairments in micturition reflex.

Estrogen modulates the spinal N-methyl-D-aspartic acid-mediated pelvic nerve-to-urethra reflex plasticity in rats.

The effects of gonadal steroids on glutamate-mediated pelvic nerve-to-urethra reflex (PUR) plasticity were investigated in rats, which received a sham operation (Sham), ovariectomy (OVX), or ovariectomy with daily supplemental estrogen (50 microg/kg, OVX + E2). The magnitude of the repetitive stimulation (RS, 1 Hz)-induced potentiation in PUR activity decreased significantly in the OVX group when compared with the Sham groups (18.09 +/- 3.91 and 7.40 +/- 1.03 spikes/stimulation in Sham and OVX group; respectively, P < 0.01, n = 21). Supplemental estrogen (OVX + E2, 12.60 +/- 1.49 spikes/stimulation) significantly reversed the decrease in RS-induced PUR potentiation caused by OVX (P < 0.01, n = 21). The magnitude of the RS-induced potentiation in PUR activity decreased significantly after intrathecal 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo (F) quinoxaline [(20 microm, 10 microl), from 18.09 +/- 3.91 to 10.40 +/- 0.81, from 7.40 +/- 1.03 to 3.20 +/- 0.94, and from 12.60 +/- 1.49 to 8.06 +/- 0.32 spikes/stimulation in Sham, OVX, and OVX + E2, respectively, P < 0.01, n = 18] and D-2-amino-5-phosphonoraleric acid [(100 microm, 10 microl), from 18.09 +/- 3.91 to 1.04 +/- 0.12, from 7.40 +/- 1.03 to 1.06 +/- 0.22, and from 12.60 +/- 1.49 to 0.98 +/- 0.25 spikes/stimulation in Sham, OVX, and OVX + E2, respectively, P < 0.01, n = 18]. In addition, potentiation in PUR activities was induced by intrathecal l-glutamate (0.1 mm, 10 microl, from 1.04 +/- 0.02 to 21.60 +/- 0.93, from 1.10 +/- 0.06 to 8.40 +/- 1.50, and from 1.03 +/- 0.03 to 18.04 +/- 0.84 spikes/stimulation in Sham, OVX, and OVX + E2, respectively, P < 0.01, n = 18) and N-methyl-D-aspartic acid (0.1 mm, 10 microl, from 1.04 +/- 0.02 to 14.80 +/- 0.97, from 1.10 +/- 0.06 to 4.60 +/- 0.48, and from 1.03 +/- 0.03 to 9.09 +/- 0.63 spikes/stimulation in Sham, OVX, and OVX + E2); N-methyl-D-aspartic acid-mediated PUR plasticity in female rats and may contribute to alterations in urinary dysfunction after menopause.

Impaired micturition reflex caused by acute selective dorsal or ventral root(s) rhizotomy in anesthetized rats.

PURPOSE: To clarify the contributions of parasympathetic inputs and outputs to the micturition reflex. MATERIALS AND METHODS: Intra-vesical pressure (IVP), external urethral sphincter electromyogram (EMG), pelvic afferent nerve activities (PANA), and pelvic efferent nerve activities (PENA) as well as the time-derived IVP (dIVP, an index of bladder contractility) were evaluated in intact and acute dorsal or ventral root(s) rhizotomized (DRX and VRX, respectively) rats. RESULTS: In DRX rats, when compared with that in intact stage, the voiding frequency was decreased (75 +/- 15% of intact, P < 0.05, n = 8), while the threshold pressure to trigger voiding contractions was significantly increased (187 +/- 75% of intact, P < 0.05, n = 8). In addition, several insufficient contractions (5.3 +/- 3.5 contractions/voiding, P < 0.05, n = 8) occurred in ahead of each voiding contraction. On the other hand, in VRX rats, the peak and rebound IVP were significantly decreased (90 +/- 3.5% and 75 +/- 11.3% of intact, P < 0.01, n = 8), while the threshold pressure was not affected (102 +/- 11% of intact, P = NS, n = 8). The time-derived parameters were significantly decreased in VRX (peak dIVP, 78 +/- 10.2%, rebound dIVP, 75 +/- 15.6%, minimal dIVP, 68 +/- 14% of intact, P < 0.01, n = 8) but only peak dIVP was decreased (85 +/- 11% of intact, P < 0.01, n = 8) in DRX rats. CONCLUSION: Acute selective DRX and VRX rat can be an animal model to investigate peripheral neural control in micturition functions.

Mitochondrial Na+ overload is caused by oxidative stress and leads to activation of the caspase 3- dependent apoptotic machinery.

Oxidative stress is one of the major causes of cell death. Using time-lapse confocal recording of live cardiomyocytes, we showed that H2O2 (OH*) caused a marked increase in Na+ and Ca2+ levels in both the cytosol ([Na]cyt, [Ca]cyt) and mitochondria ([Na]m, [Ca]m). The H2O2-induced intracellular Na+ ([Na]i) overload contributed to the H2O2-induced [Ca]cyt/[Ca]m overload via activation of the reverse mode of the Na-Ca exchanger. When myocytes were treated for 40 min with 100 microM H2O2 in normal medium, then returned to H2O2-free medium, the percentage of apoptotic cells increased from 4% at 0 h to 55 and 85% at 4.5 and 16 h, respectively. H2O2-induced apoptosis was completely prevented by using Na-free, but not Ca-free, medium. When a Na+ ionophore cocktail in Ca-free medium was used instead of H2O2 to increase the [Na]i by more than 30 mM without any change in the [Ca]i, cytochrome c release and caspase 3-dependent apoptosis occurred, showing that [Na]i overload per se induced apoptosis. We also showed that the increase in the mitochondrial, but not the cytosolic, Na+ levels resulted in the opening of the permeation transition pore, followed by cytochrome c release. Our findings therefore suggest that H2O2-induced [Na]m overload is an important upstream signal for the apoptotic machinery, and the prevention of [Na]m overload thus represents a particularly attractive target for strategies aimed at preventing oxidative stress-induced cell death.