Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 36
Filter
1.
Braz. arch. biol. technol ; 63: e20180292, 2020. tab, graf
Article in English | LILACS | ID: biblio-1132184

ABSTRACT

Abstract Acetylated cassava starch with low and medium degrees of substitution (DS) were synthesized. Also, the effect of DS on swelling power, solubility, morphological properties, gelatinization temperature, paste clarity and moisture sorption were studied. Swelling power and solubility in water between 50ºC and 90°C were determined. Acetylated cassava starches with low DS showed an increased in both parameters, while at higher DS values a reduction of them was observed. Maximum swelling power values were measured in acetylated starch with DS of 0.2 and maximum solubility was registered at DS of 0.72. Equilibrium moisture content values from sorption isotherms presented a good fit using the GAB model (R2>0.96). SEM micrographs showed that as acetyl groups are incorporated the granules suffer surface changes and eventually lose their structure at DS of 1.5. Clarity of acetylated starch pastes with low DS was lighter than native starch paste. In addition, the increase in DS produced a reduction in gelatinization temperature.


Subject(s)
Acetylation , Solubility , Substantia Gelatinosa , Temperature , Starch and Fecula , Microscopy, Electron, Scanning
2.
Article in English | WPRIM | ID: wpr-764103

ABSTRACT

PURPOSE: The aim of this study was to characterize the responsiveness of miniature excitatory postsynaptic currents (mEPSCs) to α1-adrenoceptor blockers in substantia gelatinosa (SG) neurons from the spinal cord to develop an explanation for the efficacy of α1-adrenoceptor blockers in micturition dysfunction. METHODS: Male adult Sprague-Dawley rats were used. Blind whole-cell patch-clamp recordings were performed using SG neurons in spinal cord slices. Naftopidil (100μM), tamsulosin (100μM), or silodosin (30μM), α1-adrenoceptor blockers, was perfused. The frequency of mEPSCs was recorded in an SG neuron to which the 3 blockers were applied sequentially with wash-out periods. Individual frequencies in a pair before naftopidil and tamsulosin perfusion were plotted as baseline, and the correlation between them was confirmed by Spearman correlation coefficient; linear regression was then performed. The same procedure was performed before naftopidil and silodosin perfusion. Frequencies of pairs after naftopidil and tamsulosin perfusion and after naftopidil and silodosin perfusion were similarly analyzed. The ratios of the frequencies after treatment to before were then calculated. RESULTS: After the treatments, Spearman ρ and the slope were decreased to 0.682 from 0.899 at baseline and 0.469 from 1.004 at baseline, respectively, in the tamsulosin group relative to the naftopidil group. In the silodosin group, Spearman ρ and the slope were also decreased to 0.659 from 0.889 at baseline and 0.305 from 0.989 at baseline, respectively, relative to the naftopidil group. Naftopidil significantly increased the ratio of the frequency of mEPSCs compared to tamsulosin and silodosin (P=0.015 and P=0.004, respectively). CONCLUSIONS: There was a difference in responsiveness in the frequency of mEPSCs to α1-adrenoceptor blockers, with the response to naftopidil being the greatest among the α1-adrenoceptor blockers. These data are helpful to understand the action mechanisms of α1-adrenoceptor blockers for male lower urinary tract symptoms in clinical usage.


Subject(s)
Adrenergic alpha-1 Receptor Antagonists , Adult , Animals , Excitatory Postsynaptic Potentials , Humans , Linear Models , Lower Urinary Tract Symptoms , Male , Neurons , Perfusion , Rats , Rats, Sprague-Dawley , Spinal Cord , Substantia Gelatinosa , Urination
3.
Article in English | WPRIM | ID: wpr-761789

ABSTRACT

The lamina II, also called the substantia gelatinosa (SG), of the trigeminal subnucleus caudalis (Vc), is thought to play an essential role in the control of orofacial nociception. Glycine and serotonin (5-hydroxytryptamine, 5-HT) are the important neurotransmitters that have the individual parts on the modulation of nociceptive transmission. However, the electrophysiological effects of 5-HT on the glycine receptors on SG neurons of the Vc have not been well studied yet. For this reason, we applied the whole-cell patch clamp technique to explore the interaction of intracellular signal transduction between 5-HT and the glycine receptors on SG neurons of the Vc in mice. In nine of 13 neurons tested (69.2%), pretreatment with 5-HT potentiated glycine-induced current (I(Gly)). Firstly, we examined with a 5-HT₁ receptor agonist (8-OH-DPAT, 5-HT(1/7) agonist, co-applied with SB-269970, 5-HT₇ antagonist) and antagonist (WAY-100635), but 5-HT₁ receptor agonist did not increase IGly and in the presence of 5-HT₁ antagonist, the potentiation of 5-HT on I(Gly) still happened. However, an agonist (α-methyl-5-HT) and antagonist (ketanserin) of the 5-HT₂ receptor mimicked and inhibited the enhancing effect of 5-HT on I(Gly) in the SG neurons, respectively. We also verified the role of the 5-HT₇ receptor by using a 5-HT₇ antagonist (SB-269970) but it also did not block the enhancement of 5-HT on I(Gly). Our study demonstrated that 5-HT facilitated I(Gly) in the SG neurons of the Vc through the 5-HT₂ receptor. The interaction between 5-HT and glycine appears to have a significant role in modulating the transmission of the nociceptive pathway.


Subject(s)
Animals , Glycine , Mice , Neurons , Neurotransmitter Agents , Nociception , Patch-Clamp Techniques , Receptors, Glycine , Serotonin , Signal Transduction , Substantia Gelatinosa
4.
Article in English | WPRIM | ID: wpr-727870

ABSTRACT

Botulinum toxin type A (BoNT/A) has been used therapeutically for various conditions including dystonia, cerebral palsy, wrinkle, hyperhidrosis and pain control. The substantia gelatinosa (SG) neurons of the trigeminal subnucleus caudalis (Vc) receive orofacial nociceptive information from primary afferents and transmit the information to higher brain center. Although many studies have shown the analgesic effects of BoNT/A, the effects of BoNT/A at the central nervous system and the action mechanism are not well understood. Therefore, the effects of BoNT/A on the spontaneous postsynaptic currents (sPSCs) in the SG neurons were investigated. In whole cell voltage clamp mode, the frequency of sPSCs was increased in 18 (37.5%) neurons, decreased in 5 (10.4%) neurons and not affected in 25 (52.1%) of 48 neurons tested by BoNT/A (3 nM). Similar proportions of frequency variation of sPSCs were observed in 1 and 10 nM BoNT/A and no significant differences were observed in the relative mean frequencies of sPSCs among 1–10 nM BoNT/A. BoNT/A-induced frequency increase of sPSCs was not affected by pretreated tetrodotoxin (0.5 µM). In addition, the frequency of sIPSCs in the presence of CNQX (10 µM) and AP5 (20 µM) was increased in 10 (53%) neurons, decreased in 1 (5%) neuron and not affected in 8 (42%) of 19 neurons tested by BoNT/A (3 nM). These results demonstrate that BoNT/A increases the frequency of sIPSCs on SG neurons of the Vc at least partly and can provide an evidence for rapid action of BoNT/A at the central nervous system.


Subject(s)
6-Cyano-7-nitroquinoxaline-2,3-dione , Animals , Botulinum Toxins , Botulinum Toxins, Type A , Brain , Central Nervous System , Cerebral Palsy , Dystonia , Hyperhidrosis , Mice , Neurons , Substantia Gelatinosa , Synaptic Potentials , Tetrodotoxin
5.
Article in English | WPRIM | ID: wpr-727577

ABSTRACT

The superficial dorsal horn of the spinal cord plays an important role in pain transmission and opioid activity. Several studies have demonstrated that opioids modulate pain transmission, and the activation of µ-opioid receptors (MORs) by opioids contributes to analgesic effects in the spinal cord. However, the effect of the activation of MORs on GABAergic interneurons and the contribution to the analgesic effect are much less clear. In this study, using transgenic mice, which allow the identification of GABAergic interneurons, we investigated how the activation of MORs affects the excitability of GABAergic interneurons and synaptic transmission between primary nociceptive afferent and GABAergic interneurons. We found that a selective µ-opioid agonist, [D-Ala², NMe-Phe⁴, Gly-ol]-enkephanlin (DAMGO), induced an outward current mediated by K⁺ channels in GABAergic interneurons. In addition, DAMGO reduced the amplitude of evoked excitatory postsynaptic currents (EPSCs) of GABAergic interneurons which receive monosynaptic inputs from primary nociceptive C fibers. Taken together, we found that DAMGO reduced the excitability of GABAergic interneurons and synaptic transmission between primary nociceptive C fibers and GABAergic interneurons. These results suggest one possibility that suppression of GABAergic interneurons by DMAGO may reduce the inhibition on secondary GABAergic interneurons, which increase the inhibition of the secondary GABAergic interneurons to excitatory neurons in the spinal dorsal horn. In this circumstance, the sum of excitation of the entire spinal network will control the pain transmission.


Subject(s)
Analgesics, Opioid , Animals , Enkephalin, Ala(2)-MePhe(4)-Gly(5)- , Excitatory Postsynaptic Potentials , GABAergic Neurons , Interneurons , Mice , Mice, Transgenic , Nerve Fibers, Unmyelinated , Neurons , Spinal Cord , Spinal Cord Dorsal Horn , Substantia Gelatinosa , Synaptic Transmission
6.
Article in Korean | WPRIM | ID: wpr-740082

ABSTRACT

Reactive oxygen species (ROS) and nitrogen species (RNS) are involved in cellular signaling processes as a cause of oxidative stress. According to recent studies, ROS and RNS are important signaling molecules involved in pain transmission through spinal mechanisms. In this study, a patch clamp recording was used in spinal slices of rats to investigate the action mechanisms of O₂˙⁻ and NO on the excitability of substantia gelatinosa (SG) neuron. The application of xanthine and xanthine oxidase (X/XO) compound, a ROS donor, induced inward currents and increased the frequency of spontaneous excitatory postsynaptic currents (sEPSC) in slice preparation. The application of S-nitroso-N-acetyl-DLpenicillamine (SNAP), a RNS donor, also induced inward currents and increased the frequency of sEPSC. In a single cell preparation, X/XO and SNAP had no effect on the inward currents, revealing the involvement of presynaptic action. X/XO and SNAP induced a membrane depolarization in current clamp conditions which was significantly decreased by the addition of thapsigargin to an external calcium free solution for blocking synaptic transmission. Furthermore, X/XO and SNAP increased the frequency of action potentials evoked by depolarizing current pulses, suggesting the involvement of postsynaptic action. According to these results, it was estblished that elevated ROS and RNS in the spinal cord can sensitize the dorsal horn neurons via pre- and postsynaptic mechanisms. Therefore, ROS and RNS play similar roles in the regulation of the membrane excitability of SG neurons.


Subject(s)
Action Potentials , Animals , Calcium , Excitatory Postsynaptic Potentials , Humans , Membranes , Neurons , Nitric Oxide , Nitrogen , Oxidative Stress , Posterior Horn Cells , Rats , Reactive Oxygen Species , Spinal Cord , Substantia Gelatinosa , Superoxides , Synaptic Transmission , Thapsigargin , Tissue Donors , Xanthine , Xanthine Oxidase
7.
Article in English | WPRIM | ID: wpr-718570

ABSTRACT

PURPOSE: Naftopidil ((±)-1-[4-(2-methoxyphenyl) piperazinyl]-3-(1-naphthyloxy) propan-2-ol) is prescribed in several Asian countries for lower urinary tract symptoms suggestive of benign prostatic hyperplasia. Previous animal experiments showed that intrathecal injection of naftopidil abolished rhythmic bladder contraction in vivo. Naftopidil facilitated spontaneous inhibitory postsynaptic currents in substantia gelatinosa (SG) neurons in spinal cord slices. These results suggest that naftopidil may suppress the micturition reflex at the spinal cord level. However, the effect of naftopidil on evoked excitatory postsynaptic currents (EPSCs) in SG neurons remains to be elucidated. METHODS: Male Sprague-Dawley rats at 6 to 8 weeks old were used. Whole-cell patch-clamp recordings were made using SG neurons in spinal cord slices isolated from adult rats. Evoked EPSCs were analyzed in Aδ or C fibers. Naftopidil or prazosin, an α1-adrenoceptor blocker, was perfused at 100 μM or 10 μM, respectively. RESULTS: Bath-applied 100 μM naftopidil significantly decreased the peak amplitudes of Aδ and C fiber-evoked EPSCs to 72.0%±7.1% (n=15) and 70.0%±5.5% (n=20), respectively, in a reversible and reproducible manner. Bath application of 10μM prazosin did not inhibit Aδ or C fiber-evoked EPSCs. CONCLUSIONS: The present study suggests that a high concentration of naftopidil reduces the amplitude of evoked EPSCs via a mechanism that apparently does not involve α1-adrenoceptors. Inhibition of evoked EPSCs may also contribute to suppression of the micturition reflex, together with nociceptive stimulation.


Subject(s)
Adult , Animal Experimentation , Animals , Asian Continental Ancestry Group , Baths , Excitatory Postsynaptic Potentials , Humans , In Vitro Techniques , Inhibitory Postsynaptic Potentials , Injections, Spinal , Lower Urinary Tract Symptoms , Male , Nerve Fibers, Unmyelinated , Neurons , Prazosin , Prostatic Hyperplasia , Rats , Rats, Sprague-Dawley , Reflex , Spinal Cord , Substantia Gelatinosa , Urinary Bladder , Urination
8.
Article in Korean | WPRIM | ID: wpr-54240

ABSTRACT

Recent studies indicate that mitochondria are an important source of reactive oxygen species (ROS) in the spinal dorsal horn. In our previous study, application of malate, a mitochondrial electron transport complex I substrate, induced a membrane depolarization, which was inhibited by pretreatment with ROS scavengers. In the present study, we used patch clamp recording in the substantia geletinosa (SG) neurons of spinal slices, to investigate the cellular mechanism of mitochondrial ROS on neuronal excitability. DNQX (an AMPA receptor antagonist) and AP5 (an NMDA receptor antagonist) decreased the malate-induced depolarization. In an external calcium free solution and addition of tetrodotoxin (TTX) for blockade of synaptic transmission, the malateinduced depolarization remained unchanged. In the presence of DNQX, AP5 and AP3 (a group I metabotropic glutamate receptor (mGluR) antagonist), glutamate depolarized the membrane potential, which was suppressed by PBN. However, oligomycin (a mitochondrial ATP synthase inhibitor) or PPADS (a P2 receptor inhibitor) did not affect the substrates-induced depolarization. These results suggest that mitochondrial substrate-induced ROS in SG neuron directly acts on the postsynaptic neuron, therefore increasing the ion influx via glutamate receptors.


Subject(s)
Animals , Calcium , Electron Transport Complex I , Glutamic Acid , Membrane Potentials , Membranes , Mitochondria , Mitochondrial Proton-Translocating ATPases , N-Methylaspartate , Neurons , Oligomycins , Rats , Reactive Oxygen Species , Receptors, AMPA , Receptors, Glutamate , Receptors, Metabotropic Glutamate , Spinal Cord Dorsal Horn , Substantia Gelatinosa , Synaptic Transmission , Tetrodotoxin
9.
Article in English | WPRIM | ID: wpr-145728

ABSTRACT

BACKGROUND: Intravenous palonosetron-HCl, a second-generation antagonist of selective serotonin type 3 (5-HT3) receptors, can prevent chemotherapy-induced nausea and vomiting (CINV) and postoperative nausea and vomiting (PONV). 5-HT3 receptors are abundant in the lower brainstem and the substantia gelatinosa of the spinal cord, which provides a theoretical rationale for neuraxial administration of 5-HT3 receptor antagonists for CINV, PONV, and opioid-induced nausea and vomiting. However, there are no reports of neuraxial administration of palonosetron-HCl. Before neuraxial administration of a drug is accepted for clinical use, its safety must be proven. This study was conducted to determine whether neuraxial administration of palonosetron-HCl produces neurologic injury. METHODS: Male Sprague-Dawley rats under general anesthesia were catheterized intrathecally and the catheter tip was advanced caudally to the L1 vertebra. After 7 days, 20 µl of normal saline (N group, n = 6) or 20 µl (1 µg) of palonosetron-HCl (P group, n = 6) were injected intrathecally once per day for 2 weeks. Neurotoxic changes were evaluated by light microscopy (LM) and electron microscopy (EM) of the spinal cord. Behavioral changes were also evaluated in both groups. RESULTS: One of the N group rats and three of the P group rats demonstrated abnormal behavior during intrathecal drug injection, but otherwise their behavior was normal. The spinal cords of the N group did not have any abnormal findings by LM or EM. The spinal cords of the P group had multiple vacuoles in the white matter by LM, especially in the dorsal funiculus, and EM revealed myelin, axonal, and mitochondrial swelling. CONCLUSIONS: Results suggest that chronic intrathecal administration of palonosetron-HCl produced microscopic morphologic changes in the spinal cords of rats.


Subject(s)
Anesthesia, General , Animals , Axons , Brain Stem , Catheters , Humans , Injections, Spinal , Male , Microscopy , Microscopy, Electron , Mitochondrial Swelling , Myelin Sheath , Nausea , Postoperative Nausea and Vomiting , Rats , Rats, Sprague-Dawley , Receptors, Serotonin, 5-HT3 , Serotonin , Spinal Cord , Spine , Substantia Gelatinosa , Vacuoles , Vomiting , White Matter
10.
J. appl. oral sci ; 24(3): 218-222, graf
Article in English | LILACS, BBO | ID: lil-787544

ABSTRACT

ABSTRACT The most recently identified serotonin (5-HT) receptor is the 5-HT7 receptor. The antinociceptive effects of a 5-HT7 receptor agonist have been shown in neuropathic and inflammatory animal models of pain. A recent study demonstrated the functional expression of 5-HT7 receptors in the substantia gelatinosa (SG) of the trigeminal subnucleus caudalis, which receives and processes orofacial nociceptive inputs. Objective To investigate the antinociceptive effects of pharmacological activation of 5-HT7 receptors on orofacial pain in mice. Material and Methods Nociception was evaluated by using an orofacial formalin test in male Balb-C mice. Selective 5-HT7 receptor agonists, LP 44 and LP 211 (1, 5, and 10 mg/kg), were given intraperitoneally 30 min prior to a formalin injection. A bolus of 10 µl of 4% subcutaneous formalin was injected into the upper lip of mice and facial grooming behaviors were monitored. The behavioral responses consisted of two distinct periods, the early phase corresponding to acute pain (Phase I: 0–12 min) and the late phase (Phase II: 12–30 min). Results LP 44 and LP 211 (1, 5, and 10 mg/kg) produced an analgesic effect with reductions in face rubbing time in both Phase I and Phase II of the formalin test. Conclusion Our results suggest that 5-HT7 receptor agonists may be promising analgesic drugs in the treatment of orofacial pain.


Subject(s)
Animals , Male , Mice , Piperazines/therapeutic use , Facial Pain/drug therapy , Receptors, Serotonin , Serotonin Receptor Agonists/therapeutic use , Analgesics/therapeutic use , Substantia Gelatinosa/drug effects , Time Factors , Trigeminal Nerve/drug effects , Facial Pain/chemically induced , Reproducibility of Results , Treatment Outcome , Disease Models, Animal , Dose-Response Relationship, Drug , Formaldehyde , Mice, Inbred BALB C
11.
Article in English | WPRIM | ID: wpr-728678

ABSTRACT

The analgesic mechanism of opioids is known to decrease the excitability of substantia gelatinosa (SG) neurons receiving the synaptic inputs from primary nociceptive afferent fiber by increasing inwardly rectifying K⁺ current. In this study, we examined whether a µ-opioid agonist, [D-Ala2,N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO), affects the two-pore domain K⁺ channel (K2P) current in rat SG neurons using a slice whole-cell patch clamp technique. Also we confirmed which subtypes of K2P channels were associated with DAMGO-induced currents, measuring the expression of K2P channel in whole spinal cord and SG region. DAMGO caused a robust hyperpolarization and outward current in the SG neurons, which developed almost instantaneously and did not show any time-dependent inactivation. Half of the SG neurons exhibited a linear I~V relationship of the DAMGO-induced current, whereas rest of the neurons displayed inward rectification. In SG neurons with a linear I~V relationship of DAMGO-induced current, the reversal potential was close to the K⁺ equilibrium potentials. The mRNA expression of TWIK (tandem of pore domains in a weak inwardly rectifying K⁺ channel) related acid-sensitive K⁺ channel (TASK) 1 and 3 was found in the SG region and a low pH (6.4) significantly blocked the DAMGO-induced K⁺ current. Taken together, the DAMGO-induced hyperpolarization at resting membrane potential and subsequent decrease in excitability of SG neurons can be carried by the two-pore domain K⁺ channel (TASK1 and 3) in addition to inwardly rectifying K⁺ channel.


Subject(s)
Analgesics, Opioid , Animals , Enkephalin, Ala(2)-MePhe(4)-Gly(5)- , Hydrogen-Ion Concentration , Membrane Potentials , Neurons , Rats , RNA, Messenger , Spinal Cord , Substantia Gelatinosa
12.
Article in Korean | WPRIM | ID: wpr-124488

ABSTRACT

Reactive oxygen species (ROS) and nitrogen species (RNS) are both important signaling molecules involved in pain transmission in the dorsal horn of the spinal cord. Xanthine oxidase (XO) is a well-known enzyme for the generation of superoxide anions (O₂˙⁻), while S-nitroso-N-acetyl-DL-penicillamine (SNAP) is a representative nitric oxide (NO) donor. In this study, we used patch clamp recording in spinal slices of rats to investigate the effects of O₂˙⁻ and NO on the excitability of substantia gelatinosa (SG) neurons. We also used confocal scanning laser microscopy to measure XO- and SNAP-induced ROS and RNS production in live slices. We observed that the ROS level increased during the perfusion of xanthine and xanthine oxidase (X/XO) compound and SNAP after the loading of 2',7'-dichlorofluorescin diacetate (H₂DCF-DA), which is an indicator of intracellular ROS and RNS. Application of ROS donors such as X/XO, β-nicotinamide adenine dinucleotide phosphate (NADPH), and 3-morpholinosydnomimine (SIN-1) induced a membrane depolarization and inward currents. SNAP, an RNS donor, also induced membrane depolarization and inward currents. X/XO-induced inward currents were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger) and manganese(III) tetrakis(4-benzoic acid) porphyrin (MnTBAP; superoxide dismutase mimetics). Nitro-L-arginine methyl ester (NAME; NO scavenger) also slightly decreased X/XO-induced inward currents, suggesting that X/XO-induced responses can be involved in the generation of peroxynitrite (ONOO⁻). Our data suggest that elevated ROS, especially O₂˙⁻, NO and ONOO⁻, in the spinal cord can increase the excitability of the SG neurons related to pain transmission.


Subject(s)
Adenine , Animals , Humans , Membranes , Microscopy, Confocal , Neurons , Nitric Oxide , Nitrogen , Perfusion , Peroxynitrous Acid , Rats , Reactive Oxygen Species , Spinal Cord , Spinal Cord Dorsal Horn , Substantia Gelatinosa , Superoxide Dismutase , Superoxides , Tissue Donors , Xanthine , Xanthine Oxidase
13.
Article in Chinese | WPRIM | ID: wpr-273787

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the rebound depolarization of substantia gelatinosa (SG) neurons in rat spinal dorsal horn and explore its modulatory mechanisms to provide better insights into rebound depolarization-related diseases.</p><p><b>METHODS</b>Parasagittal slices of the spinal cord were prepared from 3- to 5-week-old Sprague-Dawley rats. The electrophysiologic characteristics and responses to hyperpolarization stimulation were recorded using whole-cell patch-clamp technique. The effects of hyperpolarization-activated cyclic nucleotide gated cation (HCN) channel blockers and T-type calcium channel blockers on rebound depolarization of the neurons were studied.</p><p><b>RESULTS</b>A total of 63 SG neurons were recorded. Among them, 23 neurons showed no rebound depolarization, 19 neurons showed rebound depolarization without spikes, and 21 neurons showed rebound depolarization with spikes. The action potential thresholds of the neurons without rebound depolarization were significantly higher than those of the neurons with rebound depolarization and spikes (-28.7∓1.6 mV vs -36.0∓2.0 mV, P<0.05). The two HCN channel blockers CsCl and ZD7288 significantly delayed the latency of rebound depolarization with spike from 45.9∓11.6 ms to 121.6∓51.3 ms (P<0.05) and from 36.2∓10.3 ms to 73.6∓13.6 ms (P<0.05), respectively. ZD7288 also significantly prolonged the latency of rebound depolarization without spike from 71.9∓35.1 ms to 267.0∓68.8 ms (P<0.05). The T-type calcium channel blockers NiCl2 and mibefradil strongly decreased the amplitude of rebound depolarization with spike from 19.9∓6.3 mV to 9.5∓4.5 mV (P<0.05) and from 26.1∓9.4 mV to 15.5∓5.0 mV (P<0.05), respectively. Mibefradil also significantly decreased the amplitude of rebound depolarization without spike from 14.3∓3.0 mV to 7.9∓2.0 mV (P<0.05).</p><p><b>CONCLUSION</b>Nearly two-thirds of the SG neurons have rebound depolarizations modulated by HCN channel and T-type calcium channel.</p>


Subject(s)
Action Potentials , Animals , Calcium Channel Blockers , Pharmacology , Calcium Channels, T-Type , Cell Polarity , Cesium , Pharmacology , Chlorides , Pharmacology , Cyclic Nucleotide-Gated Cation Channels , Neurons , Cell Biology , Patch-Clamp Techniques , Pyrimidines , Pharmacology , Rats , Rats, Sprague-Dawley , Spinal Cord Dorsal Horn , Cell Biology , Substantia Gelatinosa , Cell Biology
14.
Braz. j. pharm. sci ; 51(1): 77-83, Jan-Mar/2015. graf
Article in English | LILACS | ID: lil-751354

ABSTRACT

The objective of this study was to investigate the formation and synthetic mechanism of related substance G in potassium clavulanate production. The impurity in the potassium clavulanate final product, with a retention time of 13.5 min, was confirmed as related substance G by high performance liquid chromatography-mass spectrometry/mass spectrometry. Related substance G analysis during the production of clavulanic acid showed that this impurity could be synthesized during fermentation, and the amount increased with the fermentation time. Studies on its synthetic mechanism showed that L-tyrosine and succinic acid were the precursors for biosynthesis of related substance G in vivo. The reaction was deduced to be catalyzed by an enzyme. The enzyme was a type of extracellular enzyme present in the fermentation supernatant.


O objetivo deste estudo foi investigar a formação e o mecanismo sintético da substância G relacionada na produção de clavulanato de potássio. A impureza do produto final clavulanato de potássio, com tempo de retenção de 13,5 min, foi confirmada como substância G relacionada por cromatografia líquida de alta eficiência-espectrometria de massas/espectrometria de massas. A análise da substância G relacionada durante a produção do ácido clavulânico mostrou que essa impureza poderia ser sintetizada durante a fermentação e que a quantidade aumenta com o tempo de fermentação. Estudos do seu mecanismo sintético mostraram que a L-tirosina e o ácido succínico foram os precursores in vivo para a biossíntese da substância G relacionada. Deduziu-se que a reação foi catalisada por uma enzima. A enzima foi do tipo extracelular, presente no sobrenadante da fermentação.


Subject(s)
Substantia Gelatinosa , Chromatography, Liquid , Clavulanic Acid , Mass Spectrometry/methods
15.
Article in Korean | WPRIM | ID: wpr-42181

ABSTRACT

Nitric Oxide (NO) is an important signaling molecule in the nociceptive process. Our previous study suggested that high concentrations of sodium nitroprusside (SNP), a NO donor, induce a membrane hyperpolarization and outward current through large conductances calcium-activated potassium (BKca) channels in substantia gelatinosa (SG) neurons. In this study, patch clamp recording in spinal slices was used to investigate the sources of Ca2+ that induces Ca2+-activated potassium currents. Application of SNP induced a membrane hyperpolarization, which was significantly inhibited by hemoglobin and 2-(4-carboxyphenyl) -4,4,5,5- tetramethylimidazoline-1-oxyl-3-oxide potassium salt (c-PTIO), NO scavengers. SNP-induced hyperpolarization was decreased in the presence of charybdotoxin, a selective BKCa channel blocker. In addition, SNP-induced response was significantly blocked by pretreatment of thapsigargin which can remove Ca2+ in endoplasmic reticulum, and decreased by pretreatment of dentrolene, a ryanodine receptors (RyR) blocker. These data suggested that NO induces a membrane hyperpolarization through BKca channels, which are activated by intracellular Ca2+ increase via activation of RyR of Ca2+ stores.


Subject(s)
Animals , Calcium , Charybdotoxin , Endoplasmic Reticulum , Humans , Membranes , Neurons , Nitric Oxide , Nitroprusside , Potassium , Rats , Ryanodine Receptor Calcium Release Channel , Ryanodine , Substantia Gelatinosa , Thapsigargin , Tissue Donors
16.
Article in Chinese | WPRIM | ID: wpr-333665

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of minocycline on hyperpolarization-activated current (Ih) in the substantia gelatinosa (SG) neurons in rat spinal dorsal horn.</p><p><b>METHODS</b>In vitro spinal cord transverse slices were prepared from 3-5-week-old male Sprague-Dawley rats. Using whole-cell patch clamp technique, Ih currents were recorded before and after bath application of minocycline (1-300 µmol/L) to the SG neurons.</p><p><b>RESULTS</b>Ih currents were observed in nearly 50% of the recorded neurons, and were blocked by Ih blocker CsCl and ZD7288. Minocycline rapidly and reversibly reduced the amplitude of Ih and decreased the current density in a concentration-dependent manner with an IC50 of 34 µmol/L.</p><p><b>CONCLUSION</b>Minocycline suppresses the excitability of SG neurons through inhibiting the amplitude and current density of Ih and thereby contributes to pain modulation.</p>


Subject(s)
Animals , Male , Minocycline , Pharmacology , Neurons , Patch-Clamp Techniques , Rats , Rats, Sprague-Dawley , Substantia Gelatinosa , Cell Biology
17.
Article in Korean | WPRIM | ID: wpr-116979

ABSTRACT

Reactive oxygen species (ROS) and nitrogen species (RNS) are implicated in cellular signaling processes and as a cause of oxidative stress. Recent studies indicate that ROS and RNS are important signaling molecules involved in nociceptive transmission. Xanthine oxidase (XO) system is a well-known system for superoxide anions (O2(.-)) generation, and sodium nitroprusside (SNP) is a representative nitric oxide (NO) donor. Patch clamp recording in spinal slices was used to investigate the role of O2(.-) and NO on substantia gelatinosa (SG) neuronal excitability. Application of xanthine and xanthine oxidase (X/XO) compound induced membrane depolarization. Low concentration SNP (10 microM) induced depolarization of the membrane, whereas high concentration SNP (1 mM) evoked membrane hyperpolarization. These responses were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger). Addition of thapsigargin to an external calcium free solution for blocking synaptic transmission, led to significantly decreased X/XO-induced responses. Additionally, X/XO and SNP-induced responses were unchanged in the presence of intracellular applied PBN, indicative of the involvement of presynaptic action. Inclusion of GDP-beta-S or suramin (G protein inhibitors) in the patch pipette decreased SNP-induced responses, whereas it failed to decrease X/XO-induced responses. Pretreatment with n-ethylmaleimide (NEM; thiol-alkylating agent) decreased the effects of SNP, suggesting that these responses were mediated by direct oxidation of channel protein, whereas X/XO-induced responses were unchanged. These data suggested that ROS and RNS play distinct roles in the regulation of the membrane excitability of SG neurons related to the pain transmission.


Subject(s)
Animals , Calcium , Ethylmaleimide , Humans , Membranes , Neurons , Nitric Oxide , Nitrogen , Nitroprusside , Oxidative Stress , Rats , Reactive Oxygen Species , Substantia Gelatinosa , Superoxides , Suramin , Synaptic Transmission , Thapsigargin , Tissue Donors , Xanthine , Xanthine Oxidase
18.
Article in Korean | WPRIM | ID: wpr-102158

ABSTRACT

Recent studies indicate that reactive oxygen species (ROS) can act as modulators of neuronal activity, and are critically involved in persistent pain primarily through spinal mechanisms. In this study, we investigated the effects of NaOCl, a ROS donor, on neuronal excitability and the intracellular calcium concentration ([Ca2+]i) in spinal substantia gelatinosa (SG) neurons. In current clamp conditions, the application of NaOCl caused a membrane depolarization, which was inhibited by pretreatment with phenyl-N-tert-buthylnitrone (PBN), a ROS scavenger. The NaOCl-induced depolarization was not blocked however by pretreatment with dithiothreitol, a sulfhydryl-reducing agent. Confocal scanning laser microscopy was used to confirm whether NaOCl increases the intracellular ROS level. ROS-induced fluorescence intensity was found to be increased during perfusion of NaOCl after the loading of 2',7'-dichlorofluorescin diacetate (H2DCF-DA). NaOCl-induced depolarization was not blocked by pretreatment with external Ca2+ free solution or by the addition of nifedifine. However, when slices were pretreated with the Ca2+ ATPase inhibitor thapsigargin, NaOCl failed to induce membrane depolarization. In a calcium imaging technique using the Ca2+-sensitive fluorescence dye fura-2, the [Ca2+]i was found to be increased by NaOCl. These results indicate that NaOCl activates the excitability of SG neurons via the modulation of the intracellular calcium concentration, and suggest that ROS induces nociception through a central sensitization.


Subject(s)
Animals , Calcium , Calcium-Transporting ATPases , Central Nervous System Sensitization , Dithiothreitol , Fluoresceins , Fluorescence , Fura-2 , Humans , Membranes , Microscopy, Confocal , Neurons , Nociception , Perfusion , Rats , Reactive Oxygen Species , Substantia Gelatinosa , Thapsigargin , Tissue Donors
19.
Article in Korean | WPRIM | ID: wpr-45684

ABSTRACT

Recent studies indicate that reactive oxygen species (ROS) are critically involved in persistent pain primarily through spinal mechanisms, and that mitochondria are the main source of ROS in the spinal dorsal horn. To investigate whether mitochondrial ROS can induce changes in membrane excitability on spinal substantia gelatonosa (SG) neurons, we examined the effects of mitochondrial electron transport complex (ETC) substrates and inhibitors on the membrane potential of SG neurons in spinal slices. Application of ETC inhibitors, rotenone or antimycin A, resulted in a slowly developing and slight membrane depolarization in SG neurons. Also, application of both malate, a complex I substrate, and succinate, a complex II substrate, caused reversible membrane depolarization and enhanced firing activity. Changes in membrane potential after malate exposure were more prominent than succinate exposure. When slices were pretreated with ROS scavengers such as phenyl-N-tert-buthylnitrone (PBN), catalase and 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPOL), malate-induced depolarization was significantly decreased. Intracellular calcium above 100 microM increased malateinduced depolarization, witch was suppressed by cyclosporin A, a mitochondrial permeability transition (MPT) inhibitor. These results suggest that enhanced production of spinal mitochondrial ROS can induce nociception through central sensitization.


Subject(s)
Animals , Antimycin A , Calcium , Catalase , Central Nervous System Sensitization , Cyclosporine , Electron Transport , Fires , Horns , Malates , Membrane Potentials , Membranes , Mitochondria , Neurons , Nociception , Permeability , Rats , Reactive Oxygen Species , Rotenone , Substantia Gelatinosa , Succinic Acid
20.
Article in English | WPRIM | ID: wpr-728329

ABSTRACT

Shilajit, a medicine herb commonly used in Ayurveda, has been reported to contain at least 85 minerals in ionic form that act on a variety of chemical, biological, and physical stressors. The substantia gelatinosa (SG) neurons of the trigeminal subnucleus caudalis (Vc) are involved in orofacial nociceptive processing. Shilajit has been reported to be an injury and muscular pain reliever but there have been few functional studies of the effect of Shilajit on the SG neurons of the Vc. Therefore, whole cell and gramicidin-perfotrated patch clamp studies were performed to examine the action mechanism of Shilajit on the SG neurons of Vc from mouse brainstem slices. In the whole cell patch clamp mode, Shilajit induced short-lived and repeatable inward currents under the condition of a high chloride pipette solution on all the SG neurons tested. The Shilajit-induced inward currents were concentration dependent and maintained in the presence of tetrodotoxin (TTX), a voltage gated Na+ channel blocker, CNQX, a non-NMDA glutamate receptor antagonist, and AP5, an NMDA receptor antagonist. The Shilajit-induced responses were partially suppressed by picrotoxin, a GABAA receptor antagonist, and totally blocked in the presence of strychnine, a glycine receptor antagonist, however not affected by mecamylamine hydrochloride (MCH), a nicotinic acetylcholine receptor antagonist. Under the potassium gluconate pipette solution at holding potential 0 mV, Shilajit induced repeatable outward current. These results show that Shilajit has inhibitory effects on the SG neurons of Vc through chloride ion channels by activation of the glycine receptor and GABAA receptor, indicating that Shilajit contains sedating ingredients for the central nervous system. These results also suggest that Shilajit may be a potential target for modulating orofacial pain processing.


Subject(s)
6-Cyano-7-nitroquinoxaline-2,3-dione , Animals , Brain Stem , Central Nervous System , Chloride Channels , Facial Pain , Gluconates , Mecamylamine , Mice , Minerals , N-Methylaspartate , Neurons , Picrotoxin , Potassium , Receptors, Glutamate , Receptors, Glycine , Receptors, Nicotinic , Resins, Plant , Strychnine , Substantia Gelatinosa , Tetrodotoxin
SELECTION OF CITATIONS
SEARCH DETAIL