Jian-qiang Kong - Academia.edu (original) (raw)
Papers by Jian-qiang Kong
Acta pharmaceutica Sinica. B, 2017
The search of new substrates with pharmaceutical and industrial potential for biocatalysts includ... more The search of new substrates with pharmaceutical and industrial potential for biocatalysts including cytochrome P450 enzymes is always challenging. Cytochrome P450 BM3 mutant 139-3, a versatile biocatalyst, exhibited hydroxylation activities towards fatty acids and alkanes. However, there were limited reports about its hydroxylation activity towards steroids. Herein, an Escherichia coli-based whole-cell extract containing the recombinant 139-3 protein was used as the biocatalyst to screen 13 steroids. Results revealed that 139-3 was able to specifically hydroxylate androstenedione (1) at 1α-position, generating a hydroxylated steroid 1α-OH-androstenedione (1a). To investigate whether C-1α hydroxylation catalyzed by BM3 mutant 139-3 could be industrially used, an optimization of catalyzing conditions was performed. Accordingly, the BM3 mutant 139-3 enzyme was observed to display maximum activity at 37 °C, under pH 7.0 for 4 h, with 37% transformation rate. Moreover, four 139-3 varian...
Journal of Pharmacology and Experimental Therapeutics
Experiments were done in isolated, perfused mesenteric vascular beds from Sprague-Dawley rats. Bo... more Experiments were done in isolated, perfused mesenteric vascular beds from Sprague-Dawley rats. Bolus injections of norepinephrine (3-100 nmol) induced dose-dependent increases in perfusion pressure with a maximum increase greater than 100 mm Hg. In the same dose range, clonidine had no effect on perfusion pressure. In the presence of an elevated pressure caused by constant infusions of norepinephrine (6-20 microM), bolus injections of clonidine (0.1-10 nmol) or acetylcholine (0.007-7 nmol) caused dose-related decreases in perfusion pressure. Procedures which damage endothelium (brief exposure to methylene blue or reactive oxygen radicals) abolished the depressor action of acetylcholine but only moderately reduced the depressor action of clonidine. The depressor action of clonidine was not antagonized by the alpha-2 adrenoceptor antagonist, idazoxan. Acetylcholine produced depressor responses in the presence of 5-hydroxy-tryptamine or vasopressin, but clonidine did not. Dose-response curves to bolus doses of norepinephrine were shifted markedly to the right by an alpha-1 selective concentration of prazosin (1 nM) and were shifted to the right with depression of maximum by infusions of clonidine (0.3 and 1.0 microM). It is concluded that, in the mesenteric vasculature of the rat: 1) the role of alpha-2 adrenoceptors, in responses to clonidine, is minimal; 2) endothelial factors play little role, if any, in the depressor effects of clonidine and 3) clonidine has a potent ability to interfere with the alpha-1 adrenoceptor-mediated vasoconstriction induced by norepinephrine. This antagonistic action may be at the level of the receptor but could involve postreceptor steps.
Yao xue xue bao = Acta pharmaceutica Sinica
Journal of Pharmacology and Experimental Therapeutics
Because alpha-1 adrenoceptor subtypes are distributed differentially in different arteries, exper... more Because alpha-1 adrenoceptor subtypes are distributed differentially in different arteries, experiments were conducted to determine the functional contribution of these subtypes in conduit vs. resistance vessels. Concentration- or dose-response curves for norepinephrine were obtained from aortic rings, superior mesenteric artery rings or the isolated perfused mesenteric vasculature from male Sprague-Dawley rats. Frequency-response curves to transmural electrical stimulation were obtained from the latter two preparations. The effects of 5-methylurapidil (5-MU), an alpha-1a adrenoceptor antagonist, and chloroethyl-clonidine (CEC), an alpha-1b adrenoceptor antagonist, on contractile responses were determined. In artery rings, 5-MU (30 nM) had no effect on the EC50 of norepinephrine but reduced the maximum response of the mesenteric artery rings by nearly 25%. In the perfused mesenteric vasculature, however, 5-MU (30 nM) shifted the ED50 for norepinephrine about 40-fold while reducing the maximum response by 30%. 5-MU depressed the frequency-response curve in the perfused mesenteric vasculature by nearly 80%, but did not alter the response of artery rings. In aorta, pretreatment with CEC (10 microM) shifted the concentration-response curve of norepinephrine by 800-fold without effecting the maximum. In mesenteric artery rings and perfused mesenteric vasculature, CEC reduced the slope and maximum response of both frequency-response and norepinephrine dose-response curves. Responses to norepinephrine (10 microM) in the perfused mesentery were abolished by 5-MU and reduced only 25% by nifedipine. These data suggest that the density or role of alpha-1a adrenoceptors may be greater in resistance vessels than in conduit vessels.
Journal of Pharmacology and Experimental Therapeutics
Chronic treatment of guinea pigs with morphine produces subsensitivity (tolerance) of the longitu... more Chronic treatment of guinea pigs with morphine produces subsensitivity (tolerance) of the longitudinal smooth muscle-myenteric plexus preparation to a variety of inhibitory agonists (e.g., mu opioid, alpha adrenoceptor and adenosine receptor agonists) and supersensitivity (dependence) to a variety of excitatory agonists (e.g., nicotine, 5-hydroxytryptamine and potassium ions). The present investigation was to determine if these changes in sensitivity could be related to changes in electrical properties of the S and AH neurons in the myenteric plexus. S neurons from morphine-implanted animals were significantly depolarized (7 mV) relative to those from placebo-implanted animals, whereas the membrane potential of AH neurons was unchanged. Approximately 60% of S neurons were hyperpolarized by morphine. In this subset of neurons, membranes were significantly depolarized but the threshold was unchanged in morphine-implanted animals. This means that resting potentials of S neurons from tolerant preparations are closer to threshold. The hyperpolarization produced by morphine (0.1 microM) was similar in preparations from morphine- and placebo-implanted animals. Thus, the partially depolarized state of S neurons in the myenteric plexus is the cause of the subsensitivity and supersensitivity to agonists and can explain both tolerance and dependence. Changes in opioid receptors or their coupling to potassium channels do not appear to contribute to tolerance in the longitudinal smooth muscle-myenteric plexus.
Yao xue xue bao = Acta pharmaceutica Sinica
Journal of Pharmacology and Experimental Therapeutics
Frequency-responses curves for nerve stimulation and dose-response curve for norepinephrine, 5-hy... more Frequency-responses curves for nerve stimulation and dose-response curve for norepinephrine, 5-hydroxytryptamine potassium chloride, vasopressin and acetylcholine (ACh) were determined in isolated, perfused mesenteric vascular beds from young (approximately 5 weeks) spontanelouly hypertensive (SHR) and Wistar Kyoto rats. Although mean systolic blood pressure (measured by tail cuff plethysmography) was slightly higher in the SHR, this difference was not significant. Slopes and maximum responses were increased significantly for nerve stimulation and all agonists. The basal perfusion pressure was also significantly elevated in the SHR. These differences are consistent with existing evidence that structural changes occur in blood vessels of SHR at an early stage and probably precede development of hypertension. Such structural changes could therefore contribute to development of the hypertension. Cocaine (1 microM) markedly increased responses to nerve stimulation and bolus injections of norepinephrine in preparations from SHR with little or no effect on such responses in Wistar Kyoto preparations, a result consistent with the known greater density of noradrenergic nerves in SHR vasculature. In the presence of cocaine, there was unmasked a selective super-sensitivity (significantly lower ED50) to norepinephrine in the SHR. Thus SHR mesenteric vessels may possess an alteration in adrenoreceptors or their coupling to other cellular mechanisms. Responses to ACh revealed no indication of a deficient endothelial mediated relaxation. An altered media:lumen ratio of small arteries, hypernoradrenergic innervation and supersensitivity to the transmitter may contribute to development of hypertension.
Journal of Pharmacology and Experimental Therapeutics
Ouabain acutely depolarizes most types of cells through inhibition of electrogenic Na ϩ ,K ϩ pump... more Ouabain acutely depolarizes most types of cells through inhibition of electrogenic Na ϩ ,K ϩ pumping and is a useful tool with which to study conditions that affect electrogenic pumping. Intracellular recording techniques were used with neurons of the guinea pig myenteric plexus/longitudinal muscle preparation exposed to ouabain. Of 35 S neurons exposed to ouabain (1 M), 15 were hyperpolarized by 10 Ϯ 2 mV, 11 were depolarized by 8 Ϯ 2 mV and the remaining neurons had no change in membrane potential. The nonselective potassium channel antagonist tetraethylammonium chloride (TEA; 0.5 mM) alone evoked modest (Ͻ5 mV) and inconsistent changes in the resting membrane potential of S neurons. However, in the presence of TEA, the hyperpolarizing response to 1 M ouabain was eliminated, and the proportion of cells depolarized by ouabain increased from 31% to 83%. Glibenclamide (10 M) and 100 nM iberiotoxin did not change the pattern of membrane potential changes induced by 1 M ouabain. Calcium-free buffer
Yao xue xue bao = Acta pharmaceutica Sinica
Journal of Pharmacology and Experimental Therapeutics
These experiments were designed to test two hypotheses: 1) the tolerance induced by morphine pell... more These experiments were designed to test two hypotheses: 1) the tolerance induced by morphine pellet implantation in guinea pigs will result in subsensitivity of cells in the locus ceruleus (LC), not only to morphine, but to another agonist acting on a different receptor and transduction system, namely the gamma-aminobutyric acid(A) receptor agonist, muscimol; and 2) The nonspecific (heterologous) tolerance would be associated with a partial depolarization of the tolerant cells and a decrease in the contribution of electrogenic Na(+)/K(+) pumping. Extracellular recording from LC neurons in brain slices from animals implanted with either morphine or placebo pellets established that the tolerant preparations were subsensitive to both morphine and muscimol. Immunocytochemical analysis identified the alpha(3)-subunit as the primary isoform of the Na(+)/K(+) pump in the cells under investigation. Whole-cell patch clamp recording of neurons in brain slices demonstrated that, with electrode...
The American journal of physiology, 1998
During late pregnancy, the rat undergoes massive plasma volume expansion due to cumulative renal ... more During late pregnancy, the rat undergoes massive plasma volume expansion due to cumulative renal sodium retention. In the present study, conducted in virgin, mid- (days 11-13), and late-pregnant (days 18-20) rats, we measured both Na+-K+-ATPase activity (by coupled enzyme assay) and abundance of the alpha-subunits of the Na+-K+-ATPase (by Western and slot blot analyses) in renal cortex, medulla, and brain stem. Unexpectedly, Na+-K+-ATPase in renal cortex, in both stages of pregnancy, is reduced versus the virgin, consistent with our finding of a reduced quantity of the alpha1-subunit. In renal medulla, there is a small rise in activity at midterm, but there is no difference in either activity or abundance of the alpha1-subunit in late pregnancy, when renal Na retention is maximal. In brain stem, where only alpha2- and alpha3-subunits are evident, pregnancy has no impact on enzyme activity or abundance of either isoform. In conclusion, the outcome of these experiments was unexpected ...
Blood vessels, 1991
The mesenteric vasculature of Dahl salt-sensitive (DS) rats on high-salt diet is supersensitive t... more The mesenteric vasculature of Dahl salt-sensitive (DS) rats on high-salt diet is supersensitive to nerve stimulation and to norepinephrine. The current experiments were undertaken to examine whether the enhanced sensitivity to nerve stimulation is due solely to the postsynaptic supersensitivity to norepinephrine, to increased sympathetic innervation, to altered transmitter release or to the presence of another transmitter acting as a potentiator. Catecholamine content and neuropeptide Y (NPY) presence were determined in tissues from young (approximately 5 weeks old) male Dahl rats exposed to 5 days of high (7%) or low (0.45%) salt diet. Catecholamine content from mesenteric artery, renal artery, caudal artery, right atrium, aorta, vas deferens and adrenal gland was quantified by high-pressure liquid chromatography with an electrochemical detector. A strain difference, independent of diet, between young DS and Dahl salt-resistant (DR) rats was seen only in adrenal epinephrine content...
Instruments and Applications, 2001
Pediatric Research, 1992
Using an isolated salt-perfused lung model in rabbits from 1 to 21 d of age, we measured the conc... more Using an isolated salt-perfused lung model in rabbits from 1 to 21 d of age, we measured the concentration of epinephrine in the pulmonary venous drainage and the pulmonary vascular response after a single dose of intratracheal epinephrine (0.1 microgram/g body weight). Lungs from 30 rabbits were isolated, ventilated, and perfused at one of four age groups (n = 7-8 per group). After ventilation/perfusion was judged to be stable, saline control was injected into the trachea, changes in pulmonary pressure were recorded, and perfusate was collected for 45 s. After restabilization, epinephrine was injected into the trachea, changes in pulmonary vascular pressure were recorded, and perfusate was collected for 45 s x two aliquots. Perfusate epinephrine concentrations were determined by HPLC. Little epinephrine was detected in the perfusate after control over all age groups, and little vascular response was noted. There was a significant age-related increase in perfusate epinephrine concentration as well as an age-related increase in vascular response (increased PAP), with the maximum epinephrine concentration and change in PAP noted at 14-21 d [group 4 = (1.72 +/- 0.42) x 10(4) pmol/L]. Also, in rabbits less than 6 d of age, deposition of epinephrine into the pulmonary venous drainage was delayed. In the rabbit model, the concentration of epinephrine reaching the heart via pulmonary circulation after intratracheal injection is, at birth, very low, and the pulmonary vascular response is diminished. Both increase as a function of age until 14-21 d of age. These findings may have clinical importance in human neonatal resuscitation endeavors.
Journal of Neuroscience Methods, 2006
Viable neurons in brain slices are crucial for electrophysiological studies. The present study de... more Viable neurons in brain slices are crucial for electrophysiological studies. The present study describes a new method for obtaining viable cells in several regions of the central nervous system including the ventral tegmental area, the hypothalamus, the periaqueductal grey matter and the spinal cord. The essence of the method was to use a modified artificial cerebrospinal fluid (ACSF) in which all NaCl was replaced initially by equi-osmotic glycerol. This modified glycerol-based ACSF was used during slice preparation. The underlying principle for the modification is to prevent the possible acute neurotoxic effects of passive chloride entry, subsequent cell swelling and lysis. This method significantly increased the live/dead ratio in morphology compared to the normal ACSF or sucrose-base ACSF, in which NaCl was replaced by sucrose. An examination of some electrophysiological and pharmacological properties of the neurons in these preparations, by means of current-clamp and voltage-clamp recordings, revealed similar properties of those neurons obtained with the traditional ACSF method. Due to the increase in the number of viable neurons, the new ACSF increases the productivity of experiments. Based on our data, we propose that this glycerol-based solution may protect CNS neurons.
Hypertension, 1995
The perfused mesenteric vasculature of Dahl salt-sensitive rats on a high salt diet for 5 days (p... more The perfused mesenteric vasculature of Dahl salt-sensitive rats on a high salt diet for 5 days (prehypertensive or early hypertensive) is selectively supersensitive to norepinephrine. The present goal was to determine whether that supersensitivity was maintained as hypertension developed. Littermates of salt-sensitive and salt-resistant rats (Dahl Brookhaven strain) were followed on low or high salt for up to 6 weeks. Systolic blood pressure was elevated in the salt-sensitive, high salt rats after 3 or 6 weeks but not after 5 days of the diet. The perfused mesenteric vascular beds from salt-sensitive rats were supersensitive to norepinephrine and nerve stimulation but not to potassium chloride when the rats had been maintained for 5 days or 3 weeks on the high salt diet. However, responses to norepinephrine declined after 6 weeks of the high salt diet. To determine whether sustained high blood pressure has a negative effect on mesenteric vascular responses, we conducted additional experiments with perfused mesenteric vascular beds from salt-sensitive Brookhaven (high salt, 5 weeks) and Rapp (high salt, 6 weeks) animals. Both groups exhibited significant negative correlations between in vivo systolic pressure and maximal responses of mesenteric vessels to norepinephrine and potassium chloride. We suggest that sustained hypertension in Dahl rats has a negative effect on the contractility of the mesenteric arterial system that, by 5 to 6 weeks, masks the initial supersensitivity to norepinephrine. No effects of any diet on the dilating responses of the mesenteric vessels to acetylcholine were observed in any group.
Developmental Brain Research, 2000
Cerebellar Purkinje neurons of rats have been shown to exhibit a progressive increase in resting ... more Cerebellar Purkinje neurons of rats have been shown to exhibit a progressive increase in resting membrane potential as the animals develop postnatally. The magnitude of this increase was equivalent in magnitude to the increase in the depolarizing action of ouabain, consistent with a role for the Na+/K+-pump in the hyperpolarization. Ouabain binding sites in whole cerebellum also increased with age.
Journal of Visualized Experiments, 2011
A fundamental goal to both basic and clinical neuroscience is to better understand the identities... more A fundamental goal to both basic and clinical neuroscience is to better understand the identities, molecular makeup, and patterns of connectivity that are characteristic to neurons in both normal and diseased brain. Towards this, a great deal of effort has been placed on building high-resolution neuroanatomical maps 1-3 . With the expansion of molecular genetics and advances in light microscopy has come the ability to query not only neuronal morphologies, but also the molecular and cellular makeup of individual neurons and their associated networks 4 . Major advances in the ability to mark and manipulate neurons through transgenic and gene targeting technologies in the rodent now allow investigators to 'program' neuronal subsets at will 5-6 . Arguably, one of the most influential contributions to contemporary neuroscience has been the discovery and cloning of genes encoding fluorescent proteins (FPs) in marine invertebrates 7-8 , alongside their subsequent engineering to yield an ever-expanding toolbox of vital reporters 9 . Exploiting cell type-specific promoter activity to drive targeted FP expression in discrete neuronal populations now affords neuroanatomical investigation with genetic precision.
Acta pharmaceutica Sinica. B, 2017
The search of new substrates with pharmaceutical and industrial potential for biocatalysts includ... more The search of new substrates with pharmaceutical and industrial potential for biocatalysts including cytochrome P450 enzymes is always challenging. Cytochrome P450 BM3 mutant 139-3, a versatile biocatalyst, exhibited hydroxylation activities towards fatty acids and alkanes. However, there were limited reports about its hydroxylation activity towards steroids. Herein, an Escherichia coli-based whole-cell extract containing the recombinant 139-3 protein was used as the biocatalyst to screen 13 steroids. Results revealed that 139-3 was able to specifically hydroxylate androstenedione (1) at 1α-position, generating a hydroxylated steroid 1α-OH-androstenedione (1a). To investigate whether C-1α hydroxylation catalyzed by BM3 mutant 139-3 could be industrially used, an optimization of catalyzing conditions was performed. Accordingly, the BM3 mutant 139-3 enzyme was observed to display maximum activity at 37 °C, under pH 7.0 for 4 h, with 37% transformation rate. Moreover, four 139-3 varian...
Journal of Pharmacology and Experimental Therapeutics
Experiments were done in isolated, perfused mesenteric vascular beds from Sprague-Dawley rats. Bo... more Experiments were done in isolated, perfused mesenteric vascular beds from Sprague-Dawley rats. Bolus injections of norepinephrine (3-100 nmol) induced dose-dependent increases in perfusion pressure with a maximum increase greater than 100 mm Hg. In the same dose range, clonidine had no effect on perfusion pressure. In the presence of an elevated pressure caused by constant infusions of norepinephrine (6-20 microM), bolus injections of clonidine (0.1-10 nmol) or acetylcholine (0.007-7 nmol) caused dose-related decreases in perfusion pressure. Procedures which damage endothelium (brief exposure to methylene blue or reactive oxygen radicals) abolished the depressor action of acetylcholine but only moderately reduced the depressor action of clonidine. The depressor action of clonidine was not antagonized by the alpha-2 adrenoceptor antagonist, idazoxan. Acetylcholine produced depressor responses in the presence of 5-hydroxy-tryptamine or vasopressin, but clonidine did not. Dose-response curves to bolus doses of norepinephrine were shifted markedly to the right by an alpha-1 selective concentration of prazosin (1 nM) and were shifted to the right with depression of maximum by infusions of clonidine (0.3 and 1.0 microM). It is concluded that, in the mesenteric vasculature of the rat: 1) the role of alpha-2 adrenoceptors, in responses to clonidine, is minimal; 2) endothelial factors play little role, if any, in the depressor effects of clonidine and 3) clonidine has a potent ability to interfere with the alpha-1 adrenoceptor-mediated vasoconstriction induced by norepinephrine. This antagonistic action may be at the level of the receptor but could involve postreceptor steps.
Yao xue xue bao = Acta pharmaceutica Sinica
Journal of Pharmacology and Experimental Therapeutics
Because alpha-1 adrenoceptor subtypes are distributed differentially in different arteries, exper... more Because alpha-1 adrenoceptor subtypes are distributed differentially in different arteries, experiments were conducted to determine the functional contribution of these subtypes in conduit vs. resistance vessels. Concentration- or dose-response curves for norepinephrine were obtained from aortic rings, superior mesenteric artery rings or the isolated perfused mesenteric vasculature from male Sprague-Dawley rats. Frequency-response curves to transmural electrical stimulation were obtained from the latter two preparations. The effects of 5-methylurapidil (5-MU), an alpha-1a adrenoceptor antagonist, and chloroethyl-clonidine (CEC), an alpha-1b adrenoceptor antagonist, on contractile responses were determined. In artery rings, 5-MU (30 nM) had no effect on the EC50 of norepinephrine but reduced the maximum response of the mesenteric artery rings by nearly 25%. In the perfused mesenteric vasculature, however, 5-MU (30 nM) shifted the ED50 for norepinephrine about 40-fold while reducing the maximum response by 30%. 5-MU depressed the frequency-response curve in the perfused mesenteric vasculature by nearly 80%, but did not alter the response of artery rings. In aorta, pretreatment with CEC (10 microM) shifted the concentration-response curve of norepinephrine by 800-fold without effecting the maximum. In mesenteric artery rings and perfused mesenteric vasculature, CEC reduced the slope and maximum response of both frequency-response and norepinephrine dose-response curves. Responses to norepinephrine (10 microM) in the perfused mesentery were abolished by 5-MU and reduced only 25% by nifedipine. These data suggest that the density or role of alpha-1a adrenoceptors may be greater in resistance vessels than in conduit vessels.
Journal of Pharmacology and Experimental Therapeutics
Chronic treatment of guinea pigs with morphine produces subsensitivity (tolerance) of the longitu... more Chronic treatment of guinea pigs with morphine produces subsensitivity (tolerance) of the longitudinal smooth muscle-myenteric plexus preparation to a variety of inhibitory agonists (e.g., mu opioid, alpha adrenoceptor and adenosine receptor agonists) and supersensitivity (dependence) to a variety of excitatory agonists (e.g., nicotine, 5-hydroxytryptamine and potassium ions). The present investigation was to determine if these changes in sensitivity could be related to changes in electrical properties of the S and AH neurons in the myenteric plexus. S neurons from morphine-implanted animals were significantly depolarized (7 mV) relative to those from placebo-implanted animals, whereas the membrane potential of AH neurons was unchanged. Approximately 60% of S neurons were hyperpolarized by morphine. In this subset of neurons, membranes were significantly depolarized but the threshold was unchanged in morphine-implanted animals. This means that resting potentials of S neurons from tolerant preparations are closer to threshold. The hyperpolarization produced by morphine (0.1 microM) was similar in preparations from morphine- and placebo-implanted animals. Thus, the partially depolarized state of S neurons in the myenteric plexus is the cause of the subsensitivity and supersensitivity to agonists and can explain both tolerance and dependence. Changes in opioid receptors or their coupling to potassium channels do not appear to contribute to tolerance in the longitudinal smooth muscle-myenteric plexus.
Yao xue xue bao = Acta pharmaceutica Sinica
Journal of Pharmacology and Experimental Therapeutics
Frequency-responses curves for nerve stimulation and dose-response curve for norepinephrine, 5-hy... more Frequency-responses curves for nerve stimulation and dose-response curve for norepinephrine, 5-hydroxytryptamine potassium chloride, vasopressin and acetylcholine (ACh) were determined in isolated, perfused mesenteric vascular beds from young (approximately 5 weeks) spontanelouly hypertensive (SHR) and Wistar Kyoto rats. Although mean systolic blood pressure (measured by tail cuff plethysmography) was slightly higher in the SHR, this difference was not significant. Slopes and maximum responses were increased significantly for nerve stimulation and all agonists. The basal perfusion pressure was also significantly elevated in the SHR. These differences are consistent with existing evidence that structural changes occur in blood vessels of SHR at an early stage and probably precede development of hypertension. Such structural changes could therefore contribute to development of the hypertension. Cocaine (1 microM) markedly increased responses to nerve stimulation and bolus injections of norepinephrine in preparations from SHR with little or no effect on such responses in Wistar Kyoto preparations, a result consistent with the known greater density of noradrenergic nerves in SHR vasculature. In the presence of cocaine, there was unmasked a selective super-sensitivity (significantly lower ED50) to norepinephrine in the SHR. Thus SHR mesenteric vessels may possess an alteration in adrenoreceptors or their coupling to other cellular mechanisms. Responses to ACh revealed no indication of a deficient endothelial mediated relaxation. An altered media:lumen ratio of small arteries, hypernoradrenergic innervation and supersensitivity to the transmitter may contribute to development of hypertension.
Journal of Pharmacology and Experimental Therapeutics
Ouabain acutely depolarizes most types of cells through inhibition of electrogenic Na ϩ ,K ϩ pump... more Ouabain acutely depolarizes most types of cells through inhibition of electrogenic Na ϩ ,K ϩ pumping and is a useful tool with which to study conditions that affect electrogenic pumping. Intracellular recording techniques were used with neurons of the guinea pig myenteric plexus/longitudinal muscle preparation exposed to ouabain. Of 35 S neurons exposed to ouabain (1 M), 15 were hyperpolarized by 10 Ϯ 2 mV, 11 were depolarized by 8 Ϯ 2 mV and the remaining neurons had no change in membrane potential. The nonselective potassium channel antagonist tetraethylammonium chloride (TEA; 0.5 mM) alone evoked modest (Ͻ5 mV) and inconsistent changes in the resting membrane potential of S neurons. However, in the presence of TEA, the hyperpolarizing response to 1 M ouabain was eliminated, and the proportion of cells depolarized by ouabain increased from 31% to 83%. Glibenclamide (10 M) and 100 nM iberiotoxin did not change the pattern of membrane potential changes induced by 1 M ouabain. Calcium-free buffer
Yao xue xue bao = Acta pharmaceutica Sinica
Journal of Pharmacology and Experimental Therapeutics
These experiments were designed to test two hypotheses: 1) the tolerance induced by morphine pell... more These experiments were designed to test two hypotheses: 1) the tolerance induced by morphine pellet implantation in guinea pigs will result in subsensitivity of cells in the locus ceruleus (LC), not only to morphine, but to another agonist acting on a different receptor and transduction system, namely the gamma-aminobutyric acid(A) receptor agonist, muscimol; and 2) The nonspecific (heterologous) tolerance would be associated with a partial depolarization of the tolerant cells and a decrease in the contribution of electrogenic Na(+)/K(+) pumping. Extracellular recording from LC neurons in brain slices from animals implanted with either morphine or placebo pellets established that the tolerant preparations were subsensitive to both morphine and muscimol. Immunocytochemical analysis identified the alpha(3)-subunit as the primary isoform of the Na(+)/K(+) pump in the cells under investigation. Whole-cell patch clamp recording of neurons in brain slices demonstrated that, with electrode...
The American journal of physiology, 1998
During late pregnancy, the rat undergoes massive plasma volume expansion due to cumulative renal ... more During late pregnancy, the rat undergoes massive plasma volume expansion due to cumulative renal sodium retention. In the present study, conducted in virgin, mid- (days 11-13), and late-pregnant (days 18-20) rats, we measured both Na+-K+-ATPase activity (by coupled enzyme assay) and abundance of the alpha-subunits of the Na+-K+-ATPase (by Western and slot blot analyses) in renal cortex, medulla, and brain stem. Unexpectedly, Na+-K+-ATPase in renal cortex, in both stages of pregnancy, is reduced versus the virgin, consistent with our finding of a reduced quantity of the alpha1-subunit. In renal medulla, there is a small rise in activity at midterm, but there is no difference in either activity or abundance of the alpha1-subunit in late pregnancy, when renal Na retention is maximal. In brain stem, where only alpha2- and alpha3-subunits are evident, pregnancy has no impact on enzyme activity or abundance of either isoform. In conclusion, the outcome of these experiments was unexpected ...
Blood vessels, 1991
The mesenteric vasculature of Dahl salt-sensitive (DS) rats on high-salt diet is supersensitive t... more The mesenteric vasculature of Dahl salt-sensitive (DS) rats on high-salt diet is supersensitive to nerve stimulation and to norepinephrine. The current experiments were undertaken to examine whether the enhanced sensitivity to nerve stimulation is due solely to the postsynaptic supersensitivity to norepinephrine, to increased sympathetic innervation, to altered transmitter release or to the presence of another transmitter acting as a potentiator. Catecholamine content and neuropeptide Y (NPY) presence were determined in tissues from young (approximately 5 weeks old) male Dahl rats exposed to 5 days of high (7%) or low (0.45%) salt diet. Catecholamine content from mesenteric artery, renal artery, caudal artery, right atrium, aorta, vas deferens and adrenal gland was quantified by high-pressure liquid chromatography with an electrochemical detector. A strain difference, independent of diet, between young DS and Dahl salt-resistant (DR) rats was seen only in adrenal epinephrine content...
Instruments and Applications, 2001
Pediatric Research, 1992
Using an isolated salt-perfused lung model in rabbits from 1 to 21 d of age, we measured the conc... more Using an isolated salt-perfused lung model in rabbits from 1 to 21 d of age, we measured the concentration of epinephrine in the pulmonary venous drainage and the pulmonary vascular response after a single dose of intratracheal epinephrine (0.1 microgram/g body weight). Lungs from 30 rabbits were isolated, ventilated, and perfused at one of four age groups (n = 7-8 per group). After ventilation/perfusion was judged to be stable, saline control was injected into the trachea, changes in pulmonary pressure were recorded, and perfusate was collected for 45 s. After restabilization, epinephrine was injected into the trachea, changes in pulmonary vascular pressure were recorded, and perfusate was collected for 45 s x two aliquots. Perfusate epinephrine concentrations were determined by HPLC. Little epinephrine was detected in the perfusate after control over all age groups, and little vascular response was noted. There was a significant age-related increase in perfusate epinephrine concentration as well as an age-related increase in vascular response (increased PAP), with the maximum epinephrine concentration and change in PAP noted at 14-21 d [group 4 = (1.72 +/- 0.42) x 10(4) pmol/L]. Also, in rabbits less than 6 d of age, deposition of epinephrine into the pulmonary venous drainage was delayed. In the rabbit model, the concentration of epinephrine reaching the heart via pulmonary circulation after intratracheal injection is, at birth, very low, and the pulmonary vascular response is diminished. Both increase as a function of age until 14-21 d of age. These findings may have clinical importance in human neonatal resuscitation endeavors.
Journal of Neuroscience Methods, 2006
Viable neurons in brain slices are crucial for electrophysiological studies. The present study de... more Viable neurons in brain slices are crucial for electrophysiological studies. The present study describes a new method for obtaining viable cells in several regions of the central nervous system including the ventral tegmental area, the hypothalamus, the periaqueductal grey matter and the spinal cord. The essence of the method was to use a modified artificial cerebrospinal fluid (ACSF) in which all NaCl was replaced initially by equi-osmotic glycerol. This modified glycerol-based ACSF was used during slice preparation. The underlying principle for the modification is to prevent the possible acute neurotoxic effects of passive chloride entry, subsequent cell swelling and lysis. This method significantly increased the live/dead ratio in morphology compared to the normal ACSF or sucrose-base ACSF, in which NaCl was replaced by sucrose. An examination of some electrophysiological and pharmacological properties of the neurons in these preparations, by means of current-clamp and voltage-clamp recordings, revealed similar properties of those neurons obtained with the traditional ACSF method. Due to the increase in the number of viable neurons, the new ACSF increases the productivity of experiments. Based on our data, we propose that this glycerol-based solution may protect CNS neurons.
Hypertension, 1995
The perfused mesenteric vasculature of Dahl salt-sensitive rats on a high salt diet for 5 days (p... more The perfused mesenteric vasculature of Dahl salt-sensitive rats on a high salt diet for 5 days (prehypertensive or early hypertensive) is selectively supersensitive to norepinephrine. The present goal was to determine whether that supersensitivity was maintained as hypertension developed. Littermates of salt-sensitive and salt-resistant rats (Dahl Brookhaven strain) were followed on low or high salt for up to 6 weeks. Systolic blood pressure was elevated in the salt-sensitive, high salt rats after 3 or 6 weeks but not after 5 days of the diet. The perfused mesenteric vascular beds from salt-sensitive rats were supersensitive to norepinephrine and nerve stimulation but not to potassium chloride when the rats had been maintained for 5 days or 3 weeks on the high salt diet. However, responses to norepinephrine declined after 6 weeks of the high salt diet. To determine whether sustained high blood pressure has a negative effect on mesenteric vascular responses, we conducted additional experiments with perfused mesenteric vascular beds from salt-sensitive Brookhaven (high salt, 5 weeks) and Rapp (high salt, 6 weeks) animals. Both groups exhibited significant negative correlations between in vivo systolic pressure and maximal responses of mesenteric vessels to norepinephrine and potassium chloride. We suggest that sustained hypertension in Dahl rats has a negative effect on the contractility of the mesenteric arterial system that, by 5 to 6 weeks, masks the initial supersensitivity to norepinephrine. No effects of any diet on the dilating responses of the mesenteric vessels to acetylcholine were observed in any group.
Developmental Brain Research, 2000
Cerebellar Purkinje neurons of rats have been shown to exhibit a progressive increase in resting ... more Cerebellar Purkinje neurons of rats have been shown to exhibit a progressive increase in resting membrane potential as the animals develop postnatally. The magnitude of this increase was equivalent in magnitude to the increase in the depolarizing action of ouabain, consistent with a role for the Na+/K+-pump in the hyperpolarization. Ouabain binding sites in whole cerebellum also increased with age.
Journal of Visualized Experiments, 2011
A fundamental goal to both basic and clinical neuroscience is to better understand the identities... more A fundamental goal to both basic and clinical neuroscience is to better understand the identities, molecular makeup, and patterns of connectivity that are characteristic to neurons in both normal and diseased brain. Towards this, a great deal of effort has been placed on building high-resolution neuroanatomical maps 1-3 . With the expansion of molecular genetics and advances in light microscopy has come the ability to query not only neuronal morphologies, but also the molecular and cellular makeup of individual neurons and their associated networks 4 . Major advances in the ability to mark and manipulate neurons through transgenic and gene targeting technologies in the rodent now allow investigators to 'program' neuronal subsets at will 5-6 . Arguably, one of the most influential contributions to contemporary neuroscience has been the discovery and cloning of genes encoding fluorescent proteins (FPs) in marine invertebrates 7-8 , alongside their subsequent engineering to yield an ever-expanding toolbox of vital reporters 9 . Exploiting cell type-specific promoter activity to drive targeted FP expression in discrete neuronal populations now affords neuroanatomical investigation with genetic precision.