Effects of sodium metabisulfite on sodium current in acutely isolated rat hippocampal CA1 neurons

AIM: To study the effects of sodium metabisulfite (SMB), sulfur dioxide (SO2) and its derivatives in vivo, sodium bisulfite and sulfite, on Na+ currents. The effects of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) against SMB were also studied in freshly dissociated hippocampal CA1 neurons in rats. METHODS: The whole-cell patch-clamp techniques were used in the experiments. RESULTS: ① SMB increased the voltage-activated Na+ currents in a concentration-and voltage-dependent manner. The amplitudes of Na+ currents was increased (22.36±3.28) % and (65.05±5.75)% (n=10) by SMB at 2 μmol/L and 20 μmol/L, respectively. ② SMB (10 μmol/L) did not affect the activation process, but changed the inactivation process significantly. Before and after application of 10 μmol/L SMB, the half-inactivation voltage was (-82.38±0.54) mV and (-69.39±0.41) mV (n=10, P<0.01). However, the slope factor remained unchanged. ③ SOD, CAT and GPx partly inhibited the incremental effect of SMB on Na+ currents. CONCLUSION: SMB increases Na+ currents and inhibits the steady-state inactivation, which contributes to the increase in the excitibility of neuronal cells. Its mechanism may involve oxidative damage caused by sulfur-and oxygen-centered free radicals in the rat hippocampal CA1 neurons.     

Role of oxytocin in neonatal rat hippocampal neurons after hypoxic-ischemic injury

AIM: To investigate whether oxytocin has neuroprotective effects on hippocampal CA1 pyramidal neurons from neonatal rats exposed to hypoxic-ischemic brain injury and the underlying mechanisms. METHODS: An in vitro model of hypoxic-ischemic injury was used by exposing the brain slices to oxygen-glucose deprivation (OGD) solution. Acute dissociated brain slices (6~8 slices per rat) from 8 Sprague-Dawely rats of 7~10 d old were used. The slices were randomly divided into 4 groups:control group, OGD 20 min group, OGD 40 min group and OGD+oxytocin group. The effect of oxytocin on neuronal death was evaluated by TO-PRO-3 staining. Fresh brain slices from other 20 neonatal rats were divided into OGD group, OGD+oxytocin group, OGD+dVOT (oxytocin receptor antagonist)+oxytocin group, and OGD+bicucuclline (GABA_A receptor antagonist)+oxytocin group. The onset of anoxic depolarization in the hippocampal neurons treated with different drugs was recorded by whole-cell patch-clamp techniques. RESULTS: The results of TO-PRO-3 staining showed that neuronal deaths in hippocampal CA1 area were increased over the prolonged OGD time. Oxytocin significantly reduced the hypoxic-ischemic deaths. Oxytocin dramatically prolonged the onset time of anoxic depolarization after the application of OGD solution. Both dVOT and bicuculline blocked this effect. CONCLUSION: Oxytocin plays a neuroprotective role in neonatal rat hippocampal CA1 pyramidal neurons by enhancing the inhibitory synaptic transmission via oxytocin receptors. Therefore, oxytocin is useful as a candidate for neuroprotective treatment after neonatal hypoxic-ischemic brain injury.     

Effects of piperine on abnormalities of inward rectifier potassium current and ultra rapid delayed rectifier potassium current induced by hydrogen peroxide in rabbit atrial myocytes

AIM: To study the protective effect of piperine on abnormalityies of inward rectifier potassium current (I_K1) and ultra rapid delayed rectifier potassium current (I_KUr) induced by hydrogen peroxide (H₂O₂) in single rabbit atrial myocytes. METHODS: The technique of whole-cell patch-clamp was used to study the effect of H₂O₂ at concentration of 50 μmol/L on I_K1 and I_KUr in single rabbit atrial myocytes. The protective effect of pretreatment with piperine (7 μmol/L) was also observed. RESULTS: The piperine at concentration of 7 μmol/L had no significant effect on I_K1 and I_KUr and their channel dynamics. In the presence of H₂O₂ at concentration of 50 μmol/L, the peak currents of I_K1 and I_KUr reduced significantly (P<0.05).The steady-state activation curve of I_KUr was shifted right, the steady-state inactivation curve of I_KUr was shifted left, and the recovery from inactivation of I_KUr was shifted downward. The I_KUr showed frequency-dependent characteristics. Piperine at concentration of 7 μmol/L significantly alleviated the inhibitory effect of H₂O₂ on I_K1 and I_KUr (P<0.01). In addition, piperine protected against the changes of I_KUr dynamics induced by H₂O₂. CONCLUSION: Piperine alleviates the abnormalities of I_K1 and I_KUr induced by oxidative stress in atrial myocytes.     

Effects of cardiomyopeptidin on sodium current in ventricular myocytes of guinea pigs

AIM: To determine the effect of cardiomyopeptidin on sodium current (I_Na) in ventricular myocytes of guinea pigs and to explore the mechanism of cardiomyopeptidin action at the ionic channel level. METHODS: Single ventricular myocytes of guinea pigs were obtained by enzymatic dissociation method. The whole-cell patch-clamp recording technique was used to record the change of I_Na. RESULTS: Cardiomyopeptidin (1, 5, 10, 50, 100 and 500 mg/L) decreased I_Na in a dose-dependent manner. The inhibition rates were (0±1)%, (6±2)%, (10±2)%, (15±1)%, (22±9)% and (30±6)%, respectively. The time to peak (TTP) was delayed from (2.8±0.7) ms to (3.0±0.8) ms (P<0.05) by cardiomyopeptidin (50 mg/L). In the presence of cardiomyopeptidin (50 mg/L), the current density-voltage curve of I_Na was shifted and without change of its active potential, peak potential, reversal potential, and the shape of the curve. The steady activation curve, the steady inactivation curve and the steady inactivation recovery curve of I_Na were not affected. CONCLUSION: Cardiomyopeptidin inhibits the I_Na in guinea pig ventricular myocytes, which may be one of the mechanisms of its antiarrhythmic effect.     

Effects of stretch on transient outward potassium and inward rectifier potassium current in cultured neonatal rat atrial myocytes

AIM：To investigate the effects of mechanical stretch on transient outward potassium current (Ito), inward rectifier potassium current (IK1) and action potential duration(APD) of cultured neonatal rat atrial myocytes. METHODS：Neonatal rat atrial myocytes were isolated and cultured on silicone sheeting with or without stretch for 24 h. The silicone membrane area was increased by 12% in stretched group. The cells without stretch served as control. Ito, IK1 and APD were recorded by the technique of whole-cell patch clamp. RESULTS：Compared with control group, Ito density in stretched myocytes was significantly reduced ［(16±04) pA/pF vs (121±29) pA/pF, P<001］, whereas IK1 density was increased ［(-108±08) pA/pF vs (-88±09) pA/pF, P<001］. The APDs at 50% and 90% levels of repolarization (APD50 and APD90) in the stretched cells were obviously decreased than those in non-stretched cells ［(105±14) ms vs (155±24) ms, (300±28) ms vs (563±36) ms, P<001］. CONCLUSION：Stretch stimulation leads to the reduction of Ito density, the increase in IK1 density and the shortness of APD in cultured rat atrial neonatal myocytes, which may contribute to atrial electrical remodeling induced by pressure overload.     

Modulating action of substance P on H+-gated current in rat dorsal root ganglion neurons

AIM：To investigate the effects of substance P (SP) on the modulation of H+-gated current in neurons. METHODS：Whole-cell patch-clamp technique was used to record the H+-gated current in the acutely isolated rat dorsal root ganglion (DRG) neurons at pH 4.0, pH 5.0, pH 6.0, and both SP and H+. The modulating action of SP on H+-gated current in the acutely isolated rat DRG neurons was studied by intracellular dialysis. RESULTS：H+-gated currents recorded at pH 5.0 were divided into 4 types: transient inward (T-type),sustained inward (S-type),biphasic inward (B-type) and opposite (O-type). SP exhibited obvious enhancing effect on the sustained component of S-type and B-type H+-gated currents, and the enhancing current amplitude was (85.53±22.93)%. However, the enhancing effect was not blocked by the SP receptor NK1 antagonist in about 81.8% DRG neurons. The enhancing effect was blocked by nonpeptidic SP receptor antagonist in about 75% DRG neurons. SP also exhibited obvious inhibitory effect on the sustained component of S-type and B-type H+-gated currents, and the inhibitory current amplitude was (48.46±4.45)%. However, the inhibitory effect was not blocked by the SP receptor antagonist in about 88.9% DRG neurons. After intracellular dialysis, GDP-β-S could not block the modulating action of SP on H+-gated currents. CONCLUSION：There are 4 types of H+-gated currents in acutely isolated rat DRG neurons: T-type, S-type, B-type and O-type. The modulating action of SP on H+-gated current exhibits both enhancement and inhibition. The regulatory mechanism includes G-protein-coupled receptor pathway and the binding of SP to a locus of H+-gated ion channels to exert modulating action on the H+-gated current.     

Effects of PDGFRα on melanocyte apoptosis induced by hydrogen peroxide

AIM: To investigate the effects of platelet-derived growth factor receptor α (PDGFRα) on melanocyte apoptosis induced by hydrogen peroxide (H₂O₂). METHODS: Melanocyte PIGI was used as the research object. After exposed to H₂O₂ at different concentrations, the cell viability was detected by MTT assay. The PIGI cells were transfec-ted with empty vector pCMV6 or PDGFRα over-expression vector pCMV6-PDGFRα. The transfection efficiency was determined by RT-qPCR and Western blot. The effect of H₂O₂ on the viability of the PIGI cells after over-expression of PDGFRα was measured by MTT assay. The cell apoptosis was analyzed by flow cytometry. The protein levels of p38, p-p38 and cleaved caspase-3 in the cells were detected by Western blot. DCDHF-DA was used to estemate the generation of reactive oxygen species (ROS) in the cells. RESULTS: The viability of PIGI cells decreased after exposed to H₂O₂ (P<0.05), and the half maximal inhibitory concentration of H₂O₂ was 0.7 mmol/L. Transfection with PDGFRα over-expression vector successfully induced high expression of PDGFRα at mRNA and protein levels in the PIGI cells, and increased the viability of the cells with H₂O₂ treatment (P<0.05). Over-expression of PDGFRα decreased the apoptotic rate of PIGI cells treated with H₂O₂ (P<0.05), and the level of ROS in the cells (P<0.05). The protein levels of cleaved caspase-3 and p-p38 were also decreased (P<0.05). CONCLUSION: PDGFRα inhibits the apoptosis of melanocytes induced by H₂O₂, partially reverses the growth inhibition of melanocytes by H₂O₂, and decreases the ROS level. The mechanism may be related to regulating the protein levels of p-p38 and cleaved caspase-3 in the cells.     

新书推荐：《分子克隆实验指南》（第三版）

AIM: To study the effect of panax notoginseng sponins (PNS) on L-type Ca²⁺ current in isolated right ventricle myocytes from chronic hypoxic rats. METHODS: Using whole cell patch clamp recording technique,we measured I_Ca-L in isolated right ventricle myocytes which were divided into three group:control group, chronic hy-poxic group and chronic hypoxic group with PNS(100 mg·kg^-1·d^-1). RESULTS: The result showed I_Ca-L of cells from chronic hypoxic group were significantly larger than the other two groups(P＜0.05). CONCLUSION: PNS decreases L-type Ca²⁺ current of the right ventricle myocytes from chronic hypoxic rats.     

Effect of NAC on oxidant stress and apoptosis in the hippocampal CA1 region of rats exposured to chronic intermittent hypoxia

AIM: To investigate the effect of N-acetylcystein (NAC) on oxidant stress, neuron apoptosis in the hippocampal CA1 region of rats exposured to chronic intermittent hypoxia (CIH). METHODS: 30 healthy male Wistar rats were randomly divided into three groups of 10 each, a CIH group, a NAC therapeutic group and a control group. The levels of MDA and SOD were detected by colorimetric method. Immunohistochemistry was used to examine the expression of p-JNK and TUNEL was used to detect the neuron apoptosis in the hippocampal CA1 region. RESULTS: The level of MDA in NAC group were lower than that in CIH group［(1.71±0.43) μmol/g protein vs （1.37±0.26) μmol/g protein, P<0.05)］. The activity of SOD in NAC group was higher than that in CIH group［(44.94±14.01) 103 NU/g protein vs(57.66±14.07) 103 NU/g protein, P<0.05)］. The expression of p-JNK protein and the apoptotic indices ［(0.39±0.16), (0.20±0.11)］ in NAC group were significantly lower than those in CIH group ［(0.53±0.10), (0.32±0.18), all P<0.05］. CONCLUSION: NAC protects hippocampal neuron from apoptosis by suppressing the oxidant stress in the hippocampal CA1 region and inhibiting the activation of JNK signaling pathway.     

Effects of chronic hypobaric hypoxia exposure on dendritic spine morphological changes and filamin-A expression in neurons of mouse hippocampal CA1 region

AIM:To investigate the effects of hypobaric hypoxic exposure on the morphological changes of dendritic spines and the expression of filamin-A in the neurons of mouse hippocampal CA1 region. METHODS:C57BL/6 male mice (6~8-week-old) were divided into normoxia 7 d group, normoxia 14 d group, hypobaric hypoxia 7 d group and hypobaric hypoxia 14 d group. The mice in hypobaric hypoxia exposure groups were placed in a hypobaric chamber with hypobaric hypoxia exposure to simulate the plateau at an altitude of 6 000 m. Golgi staining assay was used to observe the branch number of dendrites, and the length and density of basal and apical dendritic spines in the hippocampal CA1 region. The protein expression of filamin-A in the hippocampus of the mice was determined by Western blot. The protein expression and distribution of filamin-A in the hippocampal CA1 region were detected by immunofluorescence staining. RESULTS:Compared with normoxia exposure group, no significant difference of the number of dendritic branches in the hippocampal CA1 region after hypobaric hypoxia exposure was observed. However, the length of basal spines and apical spines was increased significantly (P<0.05), and the density of basal spines and apical spines was significantly reduced after hypobaric hypoxia exposure (P<0.01). The results of Western blot showed that the protein expression of filamin-A in the hippocampus of the mice after hypobaric hypoxia exposure was lower than that in normoxia exposure group (P<0.01 or P<0.05). Immunofluorescence staining showed that the filamin-A protein was expressed in the mouse hippocampal CA1 region, and the expression level after hypobaric hypoxia exposure was lower than that in normoxia group. CONCLUSION:Chronic hypobaric hypoxia exposure affects the protein expression level of filamin-A in the mouse hippocampal CA1 region, thus leading to the morphological changes of dendritic spines in the hippocampal CA1 region.     

Effect of Astragalus injection on expression of calmodulin after hypoxia/hypoglycemia and reoxygenation in rat hippocampal neurons

AIM: To investigate the effect of Astragalus injection on the expression of calmodulin(CaM) after hypoxia/ hypoglycemia and reoxygenation in rat hippocampal neurons.METHODS: The hippocampal neurons were cultured for 8 days and divided into 4 groups: normal control group (normal control), hypoxia/hypoglycemia and reoxygenation group (model), Astragalus injection solution group (solution control) and Astragalus injection group ( Astragalus ).The cells in all groups were treated with reoxygenation and normal medium after deprived of oxygen and glucose for 30 min except normal control group.The method of immunohistochemistry was used to measure the number of caspase-3 positive neurons.The expression of CaM at mRNA and protein levels was measured at time points of 0 h, 0.5 h, 2 h, 6 h, 24 h, 48 h, 72 h and 120 h after hypoxia/hypoglycemia and reoxygenation by RT-PCR and Western blotting, respectively.RESULTS: No difference of the parameters at all time points between model group and solution control group was found.Compared with normal control group, the numbers and the percentages of caspase-3 positive cells at all time points obviously increased in model group except at 0 h and 0.5 h (P<0.05).Compared with model group, the numbers and the percentages of caspase-3 positive cells were decreased in Astragalus injection group except at 0 h and 0.5 h (P<0.05).Compared with normal control group, the protein expression of CaM in rat hippocampal neurons at all time points obviously increased in model group (P<0.05).However, the protein expression of CaM in rat hippocampal neurons at all time points obviously decreased in Astragalus injection group as compared with model group (P<0.05).Compared with normal control group, the mRNA expression of CaM in rat hippocampal neurons at all time points obviously decreased in model group (P<0.05).The mRNA expression of CaM in rat hippocampal neurons at all time points obviously increased in Astragalus injection group as compared with model group (P<0.05).CONCLUSION: Astragalus injection inhibits the protein expression of CaM, the calcium overload and the expression of caspase-3 after hypoxia/hypoglycemia and reoxygenation, thus inhibiting hippocampal neuronal apoptosis.     

Influence of mitochondrial deficiency on expression of microtubule-associated proteins in primary cultured hippocampal neurons of neonatal rats

AIM: In order to study the relationship between mitochondrial deficiency and Alzheimer's disease(AD), we used sodium azide, a specific inhibitor of cytochrome C oxidase (COX), to develop a cell model of mitochondrial complex IV deficiency and investigated the impairment of microtubules and microtubule-associated proteins. METHODS: Primary cultured hippocampal neurons of hewborn rats were exposed to sodium azidethen cell viability was measured by MTT method; cell morphology, immunofluorecence-stained cellular microtubules and microtubule-associated proteins were observed by confocal microscopy. RESULTS: Primary cultured hippocampal neurons were exposed to 8-128 mmol/L sodium azide for 3-24 h, MTT absorbance decreased dose-and time-dependently. Exposed to 64 mmol/L sodium azide for 6 h, the processes of cells retracted, synapses disappeared, axons were shortened under contrast microscope. Meanwhile, microtubles were disassembled and became disorderly, the expression of microtubule-associated proteins were also reduced especially in the processes observed by confocal microscopy. CONCLUSION: Sodium azide inhibits the assembly and polymerization of tubulin in microtubules which may be reduced by low expression of microtubule-associated proteins in nerve cells. The damage of axons induced by microtubule collapse further blocks the intercellular signal transduction and intracellular material transportation which are important causes in cell death.     

Effects of heroin at different concentrations on electrophysiological properties of rat hippocampal CA1 pyramidal neurons

AIM: To investigate changes of electrophysiological properties of rat hippocampal CA1 pyramidal neurons in perfusion with heroin at different concentrations. METHODS: The intracellular recordings were made from CA1 pyramidal neurons of isolated hippocampal slices in perfusion with 0.03-0.3 mmol·L-1 heroin. Compared with control parametes before herion perfusion, the electrophysiological data obtained from identical neurons at 15 min after acute perfusion of the different concentration heroin were analyzed. RESULTS: With the increase of the heroin concentrations in perfusion, the absolute values of neuron resting potential and threshold potential decreased (P<0.05). In 0.3 mmol·L-1 heroin perfusion group，the amplitude of action potential decreased （P<0.05）, 10%-90% risetime and 10%-90% decaytime of spike potential elongated （P<0.05， P<0.01）, and I-V relation curve indicated membran slope resistance increased in 0.1-0.3 mmol·L^-1 perfusion group （P<0.05，P<0.01）. I-F relation curve showed that neuron average firing frequency recored at 15 min after 0.03 mmol·L^-1 heroin infusion was similar to the control frequency when intracellular stimulus intensity was 0.1-1.8 nA, however, firing frequency kept the higher level （P<0.05，P<0.01） in 1.9-2.0 nA vs the control frequence. It was evident that EPSPs of pyramidal neurons were inhibited in perfusion of three concentration heroin, especially in 0.3 mmol·L^-1. CONCLUSION: These results indicate acute heroin perfusion has a significantly influence on electrical activities of hippocampus CA1 pyramidal neurons.     

Alteration of transient outward potassium current in ventricular myocytes from 1-week and 2-month infarcted rabbit hearts

AIM: To study the current density of transient outward potassium current (I_to) in cells from the epicardial zone of the 1-week and 2-month infarcted rabbit heart. METHODS: Rabbits were infarcted by ligation of the left anterior descending coronary artery, 1 week as well as 2 months later, the single ventricular myocytes were isolated enzymatically from the infracted area of 1-week infracted rabbit heart (PMI-1 week) and 2-month infracted heart (PMI-2 months), region remote from the infracted zone of 2-month infracted heart (REM-2 months) and free wall of left ventricule from noninfarcted heart (CON). I_to was recorded using whole cell patch-clamp techniques. RESULTS: Membrane capacitance of myocytes in REM-2 months group was signifitantly larger than that in CON. I_tocurrent density (at +60 mV) was significantly reduced in PMI-1 week and PMI-2 months compared with CON , P＜0.01. Nevertheless, there was an significant increase in PMI-2M compared with PMI-1W, P＜0.05. I_to current density was also significantly decreased in REM-2 months compared with CON (P＜0.05), while there was an significant increase in REM-2 months compared with PMI-2 months, P＜0.05. CONCLUSION: The results indicated that the reduction and change of I_to in infarcted rabbit heart were heterogenous. These changes may underlie the abnormally long transmembrane action potentials of these arrhymogenic surviving ventricular fibers of the infarcted heart, thus contributing to reentrant arrhythmias in the infarcted heart. By 2 months after myocardial infarction, the depressed I_to of infracted area had returned to nearly normal, suggesting the presence of reverse remodeling.     

Effects of probucol on the proliferation of rat vascular smooth muscle cells stimulated by basic fibroblast growth factor and hydrogen peroxide

AIM: To investigate the effect of probucol on proliferation of rat vascular smooth muscle cells(VSMC) stimulated by basic fibroblast growth factor (bFGF) and/or hydrogen peroxide(H₂O₂). METHODS: Effects of probucol on VSMC proliferation and DNA synthesis stimulated by bFGF and/or H₂O₂ were observed by means of MTT test, cell number count and [3H]-TdR incorporation. RESULTS: ①Probucol significantly inhibited proliferation and DNA synthesis in VSMC stimulated by bFGF and/or H₂O₂, with dosage-dependent manner. Cell number, A value and [3H]-TdR incorporation in group probucol+bFGF and group probucol+H₂O₂ were reduced by 40.0%, 39.1%, 45.5% and 46.9%, 45.0%, 39.5%, respectively, compared with group bFGF and group H₂O₂ (P＜0.05, P＜0.01, respectively). ②Pretreatment of VSMC with probucol for 24 h prior to bFGF and/or H₂O₂ stimulation exhibited significant inhibiton of VSMC proliferation and DNA synthesis, but after prestimulation by bFGF and/or H₂O₂ for 24 h, probucol had no influence on VSMC proliferation and DNA synthesis (P＞0.05). CONCLUSION: Probucol dramatically inhibited proliferation and DNA synthesis in VSMC stimulated by bFGF and/or H₂O₂, but had no inhibitory effect on the cell proliferation prestimulated by bFGF and /or H₂O₂.     

Effects of temperature and hydrogen peroxide on mycelial growth of eight Pleurotus strains

Effective use of hydrogen peroxide as a chemical sterilant in mushroom production and selection of cultivable mushroom strains for tropical conditions require knowledge of the genetic diversity in the tolerance of the strains to hydrogen peroxide and to high temperatures. Therefore, three experiments were conducted to examine the sensitivity of Pleurotus mycelium to temperature and hydrogen peroxide. In Experiment 1, eight Pleurotus strains, which included two Pleurotus sajor-caju strains, three Pleurotus ostreatus strains, Pleurotus salmoneo stramineus, Pleurotus cornicopae and Pleurotus eryngii were cultured aseptically on agar at 25, 30 and or 35 °C. In Experiment 2, the eight strains were cultured aseptically on agar at six hydrogen peroxide concentrations (0–0.1%, v/v) at 27 °C. In Experiment 3, P. sajor-caju strain 1, a fast growing strain, was cultured non-asceptically at six hydrogen peroxide concentrations (0–0.1%, v/v) at 27 °C. In Experiment 1, mycelial growth was maximal at 25–30 °C, whereas a temperature of 35 °C was detrimental to mycelial growth except in one strain. At the highest temperature tested (35 °C), the relative mycelial growth rate (% of maximum) ranged from 6 to 91%, indicating marked differences in tolerance of the strains to high temperature. In Experiment 2, the mycelial growth rate in all strains increased when hydrogen peroxide was increased from 0 to 0.001% (v/v), and then decreased with further increments in hydrogen peroxide concentration. The strains differed markedly in sensitivity to hydrogen peroxide. The hydrogen peroxide concentration associated with 50% reduction in maximum mycelial growth rate due to toxicity (EC₅₀) ranged from 0.009 to 0.045% (v/v). It was noted that P. sajor-caju strain 1 which was the most tolerant strain to high temperature was also the most tolerant to high hydrogen peroxide concentration. In Experiment 3, involving non-aseptic culture of P. sajor-caju strain 1, bacterial growth was observed at concentrations ≤0.016%, whilst the upper hydrogen peroxide concentration limit for fungal growth was 0.025% (v/v). The highest hydrogen peroxide concentrations 0.016% (v/v) and 0.025% (v/v) in which bacteria and fungi, respectively, were observed to grow were within the concentration range 0.009–0.028% (v/v) that was found in Experiment 2 to cause a 50% reduction in mycelia growth in six of the eight Pleurotus strains tested. Use of hydrogen peroxide as a chemical sterilant in conjunction with strains highly tolerant of its toxicity offers a very cheap method of producing spawn as well as the mushrooms, and opens up opportunities for poor rural people.