Ruthenium red alleviates post-resuscitation myocardial dysfunction by upregulating mitophagy through inhibition of USP33 in a cardiac arrest rat model.

Cardiac arrest (CA) remains a leading cause of death, with suboptimal survival rates despite efforts involving cardiopulmonary resuscitation and advanced life-support technology. Post-resuscitation myocardial dysfunction (PRMD) is an important determinant of patient outcomes. Myocardial ischemia/reperfusion injury underlies this dysfunction. Previous reports have shown that ruthenium red (RR) has a protective effect against cardiac ischemia-reperfusion injury; however, its precise mechanism of action in PRMD remains unclear. This study investigated the effects of RR on PRMD and analyzed its underlying mechanisms. Ventricular fibrillation was induced in rats, which were then subjected to cardiopulmonary resuscitation to establish an experimental CA model. At the onset of return of spontaneous circulation, RR (2.5 mg/kg) was administered intraperitoneally. Our study showed that RR improved myocardial function and reduced the production of oxidative stress markers such as malondialdehyde (MDA), glutathione peroxidase (GSSG), and reactive oxygen species (ROS) production. RR also helped maintain mitochondrial structure and increased ATP and GTP levels. Additionally, RR effectively attenuated myocardial apoptosis. Furthermore, we observed downregulation of proteins closely related to mitophagy, including ubiquitin-specific protease 33 (USP33) and P62, whereas LC3B (microtubule-associated protein light chain 3B) was upregulated. The upregulation of mitophagy may play a critical role in reducing myocardial injury. These results demonstrate that RR may attenuate PRMD by promoting mitophagy through the inhibition of USP33. These effects are likely mediated through diverse mechanisms, including antioxidant activity, apoptosis suppression, and preservation of mitochondrial integrity and energy metabolism. Consequently, RR has emerged as a promising therapeutic approach for addressing post-resuscitation myocardial dysfunction.

Ruthenium red: Blocker or antagonist of TRPV channels?

Ruthenium red (RR) is a widely used inhibitor of Transient Receptor Potential (TRP) cation channels and other types of ion channels. Although RR has been generally accepted to inhibit TRP channels by physically blocking the ion permeation pathway, recent structural evidence suggests that it might also function as an antagonist, inducing conformational changes in the channel upon binding that result in closure of the pore. In a recent manuscript published in EMBO Reports, Ruth A. Pumroy and collaborators solve structures of TRPV2 and TRPV5 channels in the presence and absence of activators and RR. The data sheds light on the mechanism of inhibition by RR, while also opening new questions for further investigation.

Molecular details of ruthenium red pore block in TRPV channels.

Transient receptor potential vanilloid (TRPV) channels play a critical role in calcium homeostasis, pain sensation, immunological response, and cancer progression. TRPV channels are blocked by ruthenium red (RR), a universal pore blocker for a wide array of cation channels. Here we use cryo-electron microscopy to reveal the molecular details of RR block in TRPV2 and TRPV5, members of the two TRPV subfamilies. In TRPV2 activated by 2-aminoethoxydiphenyl borate, RR is tightly coordinated in the open selectivity filter, blocking ion flow and preventing channel inactivation. In TRPV5 activated by phosphatidylinositol 4,5-bisphosphate, RR blocks the selectivity filter and closes the lower gate through an interaction with polar residues in the pore vestibule. Together, our results provide a detailed understanding of TRPV subfamily pore block, the dynamic nature of the selectivity filter and allosteric communication between the selectivity filter and lower gate.

Hydrogen peroxide attenuates refilling of intracellular calcium store in mouse pancreatic acinar cells

Intracellular calcium (Ca²⁺) oscillation is an initial event in digestive enzyme secretion of pancreatic acinar cells. Reactive oxygen species are known to be associated with a variety of oxidative stress-induced cellular disorders including pancreatitis. In this study, we investigated the effect of hydrogen peroxide (H₂O₂) on intracellular Ca²⁺ accumulation in mouse pancreatic acinar cells. Perfusion of H₂O₂ at 300 µM resulted in additional elevation of intracellular Ca²⁺ levels and termination of oscillatory Ca²⁺ signals induced by carbamylcholine (CCh) in the presence of normal extracellular Ca²⁺. Antioxidants, catalase or DTT, completely prevented H₂O₂-induced additional Ca²⁺ increase and termination of Ca²⁺ oscillation. In Ca²⁺-free medium, H₂O₂ still enhanced CCh-induced intracellular Ca²⁺ levels and thapsigargin (TG) mimicked H₂O₂-induced cytosolic Ca²⁺ increase. Furthermore, H₂O₂-induced elevation of intracellular Ca²⁺ levels was abolished under sarco/endoplasmic reticulum Ca²⁺ ATPase-inactivated condition by TG pretreatment with CCh. H₂O₂ at 300 µM failed to affect store-operated Ca²⁺ entry or Ca²⁺ extrusion through plasma membrane. Additionally, ruthenium red, a mitochondrial Ca²⁺ uniporter blocker, failed to attenuate H₂O₂-induced intracellular Ca²⁺ elevation. These results provide evidence that excessive generation of H₂O₂ in pathological conditions could accumulate intracellular Ca²⁺ by attenuating refilling of internal Ca²⁺ stores rather than by inhibiting Ca²⁺ extrusion to extracellular fluid or enhancing Ca²⁺ mobilization from extracellular medium in mouse pancreatic acinar cells.

Mitochondrial calcium uniporter inhibition attenuates mouse bone marrow-derived mast cell degranulation induced by beta-1,3-glucan

Mast cells are primary mediators of allergic inflammation. Beta-1,3-glucan (BG) protects against infection and shock by activating immune cells. Activation of the BG receptor induces an increase in intracellular Ca2+, which may induce exocytosis. However, little is known about the precise mechanisms underlying BG activation of immune cells and the possible role of mitochondria in this process. The present study examined whether BG induced mast cell degranulation, and evaluated the role of calcium transients during mast cell activation. Our investigation focused on the role of the mitochondrial calcium uniporter (MCU) in BG-induced degranulation. Black mouse (C57) bone marrow-derived mast cells were stimulated with 0.5 microg/ml BG, 100 microg/ml peptidoglycan (PGN), or 10 microM A23187 (calcium ionophore), and dynamic changes in cytosolic and mitochondrial calcium and membrane potential were monitored. BG-induced mast cell degranulation occurred in a time-dependent manner, and was significantly reduced under calcium-free conditions. Ruthenium red, a mitochondrial Ca2+ uniporter blocker, significantly reduced mast cell degranulation induced by BG, PGN, and A23187. These results suggest that the mitochondrial Ca2+ uniporter has an important regulatory role in BG-induced mast cell degranulation.

N-acetyl-L-cysteine and cysteine increase intracellular calcium concentration in human neutrophils

N-acetyl-L-cysteine (NAC) and cysteine have been implicated in a number of human neutrophils' functional responses. However, though Ca²⁺ signaling is one of the key signalings contributing to the functional responses of human neutrophils, effects of NAC and cysteine on intracellular calcium concentration ([Ca²⁺]ᵢ) in human neutrophils have not been investigated yet. Thus, this study was carried out with an objective to investigate the effects of NAC and cysteine on [Ca²⁺]ᵢ in human neutrophils. We observed that NAC (1 µM ~ 1 mM) and cysteine (10 µM ~ 1 mM) increased [Ca²⁺]ᵢ in human neutrophils in a concentration-dependent manner. In NAC pre-supplmented buffer, an additive effect on N-formyl-methionine-leucine-phenylalanine (fMLP)-induced increase in [Ca²⁺]ᵢ in human neutrophils was observed. In Ca²⁺-free buffer, NAC- and cysteine-induced [Ca²⁺]ᵢ increase in human neutrophils completely disappeared, suggesting that NAC- and cysteine-mediated increase in [Ca²⁺]ᵢ in human neutrophils occur through Ca²⁺ influx. NAC- and cysteine-induced [Ca²⁺]ᵢ increase was effectively inhibited by calcium channel inhibitors SKF96365 (10 µM) and ruthenium red (20 µM). In Na⁺-free HEPES, both NAC and cysteine induced a marked increase in [Ca²⁺]ᵢ in human neutrophils, arguing against the possibility that Na⁺-dependent intracellular uptake of NAC and cysteine is necessary for their [Ca²⁺]ᵢ increasing activity. Our results show that NAC and cysteine induce [Ca²⁺]ᵢ increase through Ca²⁺ influx in human neutrophils via SKF96365- and ruthenium red-dependent way.

When morphogenetic proteins encounter special extracellular matrix and cell-cell connections at the interface of the renal stem/progenitor cell niche / 대한해부학회지

Reciprocal exchange of morphogenetic proteins between epithelial and mesenchymal cells in a stem/progenitor cell niche results in formation of a nephron. To maintain diffusion of morphogenetic proteins, it is assumed that a close contact exists between involved cells. However, recent publications underline that both types of stem/progenitor cells are separated by a striking interface. To explore this microarchitecture in detail, neonatal rabbit kidneys were fixed in traditional glutaraldehyde (GA) solution for transmission electron microscopy. For contrast enhancing specimens were fixed in GA solution including cupromeronic blue, ruthenium red or tannic acid. To record same perspectives, embedded blocks of parenchyma were cut in exactly orientated vertical and transverse planes to lining collecting ducts. Electron microscopy of specimens fixed by traditional GA solution illustrates a spatial separation of stem/progenitor cells and an unobstrusively looking interface. In contrast, advanced fixation of specimens in GA solution including cupromeronic blue, ruthenium red and tannic acid unmasks earlier not visible extracellular matrix. In addition, projections of mesenchymal cells covered by matrix cross the interface to contact epithelial cells. Surprisingly, the end of a mesenchymal cell projection does not dangle but is enclosed in a fitting sleeve and connected via tunneling nanotubes with the plasma membrane of an epithelial cell. Regarding this complex ensemble the question is to what extent illustrated cell-cell connections and extracellular matrix are involved in communication and transmission of morphogenetic proteins during induction of a nephron.

Different uptake of gentamicin through TRPV1 and TRPV4 channels determines cochlear hair cell vulnerability

Hair cells at the base of the cochlea appear to be more susceptible to damage by the aminoglycoside gentamicin than those at the apex. However, the mechanism of base-to-apex gradient ototoxicity by gentamicin remains to be elucidated. We report here that gentamicin caused rodent cochlear hair cell damages in a time- and dose-dependent manner. Hair cells at the basal turn were more vulnerable to gentamicin than those at the apical turn. Gentamicin-conjugated Texas Red (GTTR) uptake was predominant in basal turn hair cells in neonatal rats. Transient receptor potential vanilloid 1 (TRPV1) and 4 (TRPV4) expression was confirmed in the cuticular plate, stereocilia and hair cell body of inner hair cells and outer hair cells. The involvement of TRPV1 and TRPV4 in gentamicin trafficking of hair cells was confirmed by exogenous calcium treatment and TRPV inhibitors, including gadolinium and ruthenium red, which resulted in markedly inhibited GTTR uptake and gentamicin-induced hair cell damage in rodent and zebrafish ototoxic model systems. These results indicate that the cytotoxic vulnerability of cochlear hair cells in the basal turn to gentamicin may depend on effective uptake of the drug, which was, in part, mediated by the TRPV1 and TRPV4 proteins.

Ca(2+)-induced Ca2+ Release from Internal Stores in INS-1 Rat Insulinoma Cells

The secretion of insulin from pancreatic beta-cells is triggered by the influx of Ca2+ through voltage-dependent Ca2+ channels. The resulting elevation of intracellular calcium ([Ca2+]i) triggers additional Ca2+ release from internal stores. Less well understood are the mechanisms involved in Ca2+ mobilization from internal stores after activation of Ca2+ influx. The mobilization process is known as calcium-induced calcium release (CICR). In this study, our goal was to investigate the existence of and the role of caffeine-sensitive ryanodine receptors (RyRs) in a rat pancreatic beta-cell line, INS-1 cells. To measure cytosolic and stored Ca2+, respectively, cultured INS-1 cells were loaded with fura-2/AM or furaptra/AM. [Ca2+]i was repetitively increased by caffeine stimulation in normal Ca2+ buffer. However, peak [Ca2+]i was only observed after the first caffeine stimulation in Ca2+ free buffer and this increase was markedly blocked by ruthenium red, a RyR blocker. KCl-induced elevations in [Ca2+]i were reduced by pretreatment with ruthenium red, as well as by depletion of internal Ca2+ stores using cyclopiazonic acid (CPA) or caffeine. Caffeine-induced Ca2+ mobilization ceased after the internal stores were depleted by carbamylcholine (CCh) or CPA. In permeabilized INS-1 cells, Ca2+ release from internal stores was activated by caffeine, Ca2+, or ryanodine. Furthermore, ruthenium red completely blocked the CICR response in permeabilized cells. RyRs were widely distributed throughout the intracellular compartment of INS-1 cells. These results suggest that caffeine-sensitive RyRs exist and modulate the CICR response from internal stores in INS-1 pancreatic beta-cells.

Effects of NaOCl on the Intracellular Calcium Concentration in Rat Dorsal Root Ganglion Neurons

Recent studies have implicated reactive oxygen species (ROS) as determinants of the pathological pain caused by the activation of peripheral neurons. It has not been elucidated, however, how ROS activate the primary sensory neurons in the pain pathway. In this study, calcium imaging was performed to investigate the effects of NaOCl, a ROS donor, on the intracellular calcium concentration ([Ca2+]i) in acutely dissociated dorsal root ganglion (DRG) neurons. DRG was sequentially treated with 0.2 mg/ml of both protease and thermolysin, and single neurons were then obtained by mechanical dissociation. The administration of NaOCl then caused a reversible increase in the [Ca2+]i, which was inhibited by pretreatment with phenyl-N-tert-buthylnitrone (PBN) and isoascorbate, both ROS scavengers. The NaOCl-induced [Ca2+]i increase was suppressed both in a calcium free solution and after depletion of the intracellular Ca2+ pool by thapsigargin. Additionally, this increase was predominantly blocked by pretreatment with the transient receptor potential (TRP) antagonists, ruthenium red (50 microM) and capsazepine (10 microM). Collectively, these results suggest that an increase in the intracellular calcium concentration is produced from both extracellular fluid and the intracellular calcium store, and that TRP might be involved in the sensation of pain induced by ROS.

Effects of ruthenium red on body temperature in rats with lipopolysaccharide-induced fever / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the effect of ruthenium red (RR) on the body temperature of rats with lipopolysaccharide (LPS)-induced fever and investigate the possible mechanism.</p><p><b>METHODS</b>Rat models of fever were established with lipopolysaccharide and the effects of RR at different doses were observed on the body temperature of the rats and the content of TRPV4 in the hypothalamus.</p><p><b>RESULTS</b>Compared with those in LPS group, the rats with LPS-induced fever receiving RR treatment showed a dose-dependent lowering of the body temperature. The rats with RR treatment had lower body temperature than those with saline injection. The content of TRPV4 in the saline group was significantly higher than that in RR+LPS and RR group.</p><p><b>CONCLUSIONS</b>RR inhibits LPS-induced fever in rats and regulates the hypothalamal expression of TRPV4 channels, which may participate in the maintenance of normal body temperature.</p>

Factors modulating recovery rate after intermittent tetanic fatigue in atrophic soleus / 生理学报

Fatigue occurs when the interval of intermittent tetanic contraction of skeletal muscle is shortened to a certain degree and the contractile tension declines. After fatigue, prolongation of the contraction interval can make the contractile tension recover. In atrophic soleus, the recovery rate is slower. It has been shown that a decrease in the contractile tension is caused by the inhibition of the myofibrils and sarcoplasmic reticulum Ca(2+) release channels during fatigue. So the mechanism of the recovery of contractile tension is the recovery of the inhibited myofibrils and sarcoplasmic reticulum Ca(2+) release channels. But how the inhibition affects the recovery course is still unclear. To specify the factors modulating the recovery rate after intermittent tetanic fatigue in soleus, and to seek the reasons for the decrease in recovery rate in atrophic soleus, we observed the recovery time course of different types of fatigue in isolated soleus muscle strips. The 10% or 50% decrease in the maximal tetanic contractile tention (P(0)) was defined respectively as slight or moderate fatigue. After short-term (S10P, 10 s) and long-term (L10P, 300 s) slight fatigue, the tetanic contractile tension recovered to nearly 100% P(0) at the 20th minute. In both slight fatigue groups, perfusion with 10 mumol/L of ruthenium red (an inhibitor of Ca(2+) release channels in sarcoplasmic reticulum) slowed down the recovery rate. It was suggested that slight fatigue only induced inhibition of myofibrils. After short-term (S50P, 60 s) or long-term (L50P, 300 s) moderate fatigue, the tetanic contractile tension at the 20th minute recovered to about 95% P(0) in S50P group and 90% P(0) in L50P group, respectively. The recovery rate in L50P group was significantly lower than that in S50P group. So the recovery rate after moderate fatigue was related to the tetanic contraction duration. In both moderate fatigue groups, perfusion with 5 mmol/L of caffeine (an opener of Ca(2+) release channels in sarcoplasmic reticulum) resulted in nearly 100% recovery at the 5th minute. It was suggested that moderate fatigue induced inhibition of myofibrils and sarcoplasmic reticulum Ca(2+) release channels. In 1-week tail-suspended rats, soleus muscles showed a 40% of atrophy. After slight fatigue, the tetanic contractile tension in unloaded soleus recovered to 94% P(0) in S10P group and 95% P(0) in L10P. After moderate fatigue, the tetanic contractile tension in unloaded soleus recovered to 92% P(0) in S50P and 84% P(0) in L50P at the 20th minute. There were significant decreases in all of the fatigue groups as compared with the control groups. These results suggest that both slight and moderate fatigue inhibit the myofibrils and sarcoplasmic reticulum Ca(2+) release channels in 1-week unloaded soleus, so the recovery rate after tetanic fatigue is slower than that in the control group.

Cytotoxicity of Capsaicin on Cultured Human Skin Fibroblast / 대한피부과학회지

BACKGROUND: Capsaicin has been shown to have different biologic and toxic effects, depending on non-neuronal cells and several transformed cells, however no study has been reported from cultured human skin fibroblast. OBJECTIVE: Present study was aimed to evaluate the cytotoxicity and its mechanism of capsacin on the cultured human skin fibroblast. MATERIAL AND METHOD: Normal neonatal human fibroblasts were used, and changes of cell survival were measured by MTT assay after the cells were pre-treated with growth factors, receptor antagonist, antioxidants, calcium modulators were pre-treated or co-treated with capsaicin. RESULTS: Suvival of fibroblast was significantly increased by treatment with EGF (10ng/ml), bFGF (10ng/ml), capsazepine (10M) but inhibited by cycloheximide (1g/ml). When 200 M capsaicin was added to fibroblasts, chromatin condensations were observed at 12 hours and cell survival rate was reduced to 25-50% at 24 hours. Vanilloid receptor antagonists, capsazepine and ruthenium red, did not prevent the toxic effect of capsaicin, and 10M capsazepine paradoxically rather enhanced the cytotoxicity. In contrast to bFGF (10ng/ml), EGF (10, 100ng/ml) enhanced the cytotoxicity of capsaicin. Neuropeptides, substance P (1, 10nM) and CGRP (1, 10nM), and a structural analogue to capsaicin, tyrosine (0.3-1.2mM) did not affect the cytotoxicity. However, antioxidants such as trolox (100M) and ascorbic acid (0.1, 0.3 mM) reduced the capsaicin cytotoxicity. Of calcium modulating agents, nifedifine, a Ca2+ channel blocker (10, 20M) and cyclopiazonic acid, a Ca2+-ATPase inhibitor in ER (10M) did not influence the cytotoxicity, however BAPTA/AM (10M) as a chelater for cytoplasmic free calcium ion (10M) significantly decreased capsaicin cytotoxicity. Unlike cycloheximide, z-VAD-FMK, a protein synthesis inhibitor and a non-specific caspase inhibitor, prevented the capsaicin cytotoxicity. The DNA ladder and TUNEL positive cells were observed among the capsaicin treated fibroblasts and Western blot revealed caspase-3 activity. CONCLUSION: The capsaicin-induced cytotoxicity on human skin fibroblasts is likely to suggest the mechanism of an apoptotic pathway, which can possibly be prevented by antioxidants.

Electrophysiological effects of capsaicin on spontaneous activity of rabbit atrioventricular node cells / 生理学报

To study the electrophysiological effects of capsaicin on spontaneous activity of rabbit atrioventricular (AV) node cells, parameters of action potential in AV node were recorded using intracellular microelectrode technique. Capsaicin (1-30 micromol/L) not only decreased the amplitude of action potential, maximal rate of depolarization (V(max)), velocity of diastolic (phase 4) depolarization, and rate of pacemaker firing, but also prolonged the duration of 90% repolarization of action potential (APD(90)) in a concentration-dependent manner. Both application of L-type Ca(2+) channel agonist Bay K8644 (0.5 micromol/L) and elevation of calcium concentration (5 mmol/L) in superfusate antagonized the effects of capsaicin on pacemaker cells. Pretreatment with ruthenium red (10 micromol/L), a capsaicin receptor blocker, did not affect the effects of capsaicin on AV node cells. Capsaicin exerted an inhibitory action on spontaneous activity of AV node cells in rabbits. These effects were likely due to reduction in calcium influx, but were not mediated by VR1.

Effects of low-dose capsaicin on L-type calcium current in guinea pig ventricular myocytes / 生理学报

The purpose of this study was to examine the effects of low-dose capsaicin (CAP) on L-type calcium current (I(Ca-L) ) in guinea pig ventricular myocytes and the underlying mechanism. I(Ca-L) was examined in isolated single guinea pig ventricular myocytes by using whole-cell patch clamp technique. CAP (1-25 nmol/L) increased the voltage-dependently activated peak amplitude of I(Ca-L) and downshifted the current-voltage (I-V) curve. CAP (1, 10, 25 nmol/L) increased the peak amplitude of I(Ca-L) from -9.67+/-0.7 pA/pF to -10.21+/-0.8 pA/pF (P>0.05), to -11.37+/-0.8 pA/pF and to -12.84+/-0.9 pA/pF (P<0.05), respectively. CAP 25 nmol/L shifted the steady-state activation curve of I(Ca-L) to the left and changed half activation potential (V(0.5)) from (-20.76+/-2.0) mV to (-26.71+/-3.0) mV (P<0.05), indicating that low-dose CAP may modify the voltage-dependent activation of calcium channel. Low-dose of CAP did not affect the steady-state inactivation curve of I(Ca-L) or half-recovery time of Ca(2+) channel from inactivation. Ruthenium red (RR, 10 micromol/L), a vanilloid receptor (VR1) blocker, antagonized the effects of low-dose CAP. These results suggest that low-dose CAP increases I(Ca-L) mainly by shifting its steady-state activation curve to the left. Such effects may be mediated by VR1.

Capsaicin facilitates carotid sinus baroreflex in anesthetized rats / 生理学报

The effects of capsaicin (CAP) on the carotid sinus baroreflex were studied in 30 anaesthetized rats with perfused isolated carotid sinus. The results are as follows. (1) By perfusing the isolated carotid sinus with CAP (1 micromol/L), the functional curve of the baroreflex was shifted to the left and downward, with a peak slope (PS) increasing from 0.34+/-0.01 to 0.42+/-0.01 (P<0.01), whereas the reflex decrease (RD) in mean arterial pressure was enhanced from 36.51+/-1.26 to 45.01+/-0.71 mmHg (P<0.01). Meanwhile, the threshold pressure, equilibrium pressure and saturation pressure were all significantly decreased from 70.43 +/-2.09 to 52.86 +/-2.80 mmHg (P<0.01), 95.5+/-1.71 to 87.00+/-1.58 mmHg (P<0.01) and 177.60+/-1.37 to 163.55+/-2.12 mmHg (P<0.01), respectively. Among the functional parameters of carotid baroreflex, the changes in PS and RD induced by capsaicin were dose-dependent. (2) By pretreatment with ruthenium red (RR, 100 micromol/L), an antagonist of vanilloid receptor subtype 1 (VR(1)), the above effects of CAP on carotid baroreflex were abolished. (3) The CAP-induced change in the baroreflex was also eliminated by pretreatment with glibenclamide (20 microm ol/L), a K(ATP) channel blocker. On the basis of the results, it is concluded that CAP facilitates the carotid baroreflex, an effect of which may be resulted from the opening of K(ATP) channels mediated by VR(1).

Electrophysiological effects of capsaicin on guinea pig papillary muscles / 生理学报

The cardiac electrophysiological effects of capsaicin (CAP) were examined in guinea pig papillary muscles using intracellular microelectrode technique. The results obtained are as follows: (1) the duration of action potential (APD) in normal papillary muscles was decreased by CAP (30, 60, 120 micromol/L) in a concentration-dependent manner; (2) in partially depolarized papillary muscles, 60 micromol/L CAP not only reduced APD, but also decreased the amplitude of action potential, overshoot and maximal velocity of phase 0 depolarization; (3) pretreatment with L-type Ca(2+) channel agonist Bay K8644 (0.5 micromol/L) could completely block the effects of CAP (60 micromol/L); (4) pretreatment with ruthenium red (20 micromol/L), a vanilloid receptor (VR) blocker, did not affect the actions of capsaicin on papillary muscles. All these results suggest that the effects of CAP on papillary muscles are likely due to a decrease in calcium influx which is not mediated by VR.

Intrarenal artery injection of capsaicin activates spontaneous activity of renal afferent nerve fibers / 生理学报

The effects of intrarenal artery injection of capsaicin on multi- and single-unit spontaneous discharges of renal afferent nerve fibers were investigated in anesthetized rabbits. The results obtained are as follows: (1) intrarenal artery injection of capsaicin (20, 40, and 60 nmol/kg) increased the renal afferent nerve activity (ARNA) in a dose-dependent manner with unchanged arterial pressure; (2) pretreatment with ruthenium red (40 mmol/kg), a capsaicin receptor antagonist, completely abolished the effect of capsaicin; and (3) pretreatment with a nitric oxide synthase inhibitor L-NAME (N(6)-nitro-L-arginine methylester, 0.1 mmol/kg), significantly enhanced the ARNA response to capsaicin. The results suggest that intrarenal artery injection of capsaicin can activate ARNA via capsaicin receptors in anesthetized rabbits and that nitric oxide may be involved in regulating the activity of renal sensory nerve fibers as an inhibitory neurotransmitter.

Histochemical Electron Microscopic Findings of the Distribution of the Glycosaminoglycans of Cultured Sensory Retina

PURPOSE: This study was performed to investigate the distribution of glycosaminiglycans by the activated retinal neuronal cell on the cultured retinal tissue. METHODS: The retinal tissue was obtained from the donor eyeball without the pathological findings of the retina. The dissected retinal tissue was cultured for 2 weeks in the culture media and reacted with ruthenium red dye to observe the glycosaminoglycans reaction with transmission electron microscopy. RESULTS: The histochemical reaction to ruthenium red on the cultured retina tissue was prominent on the internal limiting membrane, intercellular space between the axons and dendrites at the outer and inner plexiform layers and interphotoreceptor matrix. CONCLUSIONS: These findings suggest that GAGs may be released into the extracellular spaces between the axons and dendrites, which modulate the retinal circuits.

Histochemical Electron Microscopic Findings of Traumatic Cataract in Rabbits by Ruthenium Red

Glycosaminoglycans (GAGs)are essential components of the extracellular matrix of various tissues.They have been known to regulate cellular and extracellular environments for cell survival and differentiation in various physiologic and pathologic conditions.The present study was conducted to identify the distribution of GAGs in subepithelial cortical fibers of normal and traumatic cataractous lens of rabbits. Traumatic cataract was made by piercing the anterior lens capsule with 25-gauge sharp needle and the rabbits were killed at different time intervals (1, 2, 3 and 4 weeks).From the enucleated eyes subepithelial lens cortical fibers were obtained.A normal rabbit lens was used for the control. The specimens were stained with 0.05%ruthenium red (RR)and processed for histochemical electron microscopy. RR reactive materials were identified as fine granular or filamentous structures.In normal rabbit lens they were present mainly along the surface of lens epithelial cells, the surface of the subepithelial cortical fibers. In cataractous rabbits lens, strong RR positive reactions were observed along the surface of the lens epithelial cells and subepthelial cortical fibers as well as at the intercortical fiber spaces and even within its micro-organelles. This investigation resulted in an illustration of the ultrastructural distribution of GAGs in normal and traumatic cataractous lens of rabbit.this may suggest altered GAG distribution may closely related to the formation of lens opacity.