Narutoshi Kabashima

Optimal dialysis improves uremic pruritus

The authors analyzed data on 59 hemodialyzed patients who did not have significant disorders of calcium and phosphate metabolism and found that more than 60% suffered from disabling pruritus possibly related to chronic uremia. Both biochemical correlates of the prevalence of pruritus and dialysis efficacy calculated by urea kinetics were investigated. Significantly higher values of blood urea nitrogen and plasma beta 2-microglobulin just before the dialysis session were observed in pruritic patients with lower dialysis efficacy estimated by Kt/V urea and normalized protein catabolic rate (nPCR). After 3 months without changing the dialysis prescriptions, 16 patients with a mean Kt/V urea and a normalized protein catabolic rate (nPCR) of 1.28 and 1.22 g/kg/d, respectively, experienced significant reductions in the degree of pruritus estimated by the pruritic score, from 12.6 +/- 5.1 to 6.3 +/- 3.2. Twenty-two patients with a mean Kt/V urea and an nPCR of 1.09 and 1.01, respectively, continued to have severe pruritus (score: 12.3 +/- 4.7 to 12.7 +/- 6.4). In 9 of 22 patients with prolonged severe pruritus, dialysis efficacy was heightened with an increase in dialyzer membrane area of more than 0.3 m2. Seven of nine patients with increased dialysis prescriptions had significant reductions of the mean pruritic score, from 12.6 +/- 4.8 to 6.3 +/- 2.4, which inversely related to the significant increase of Kt/V urea from 1.05 +/- 0.25 to 1.24 +/- 0.33; among patients whose dialysis prescriptions were not changed, only one had a significant reduction in score. The authors concluded that higher dialysis efficacy with good nutritional state reduces the prevalence and degree of pruritus in hemodialyzed patients.

Inhibition of N- and P/Q-type calcium channels by postsynaptic GABAB receptor activation in rat supraoptic neurones

1 Voltage‐dependent Ca2+ currents of dissociated rat supraoptic nucleus (SON) neurones were measured using the whole‐cell configuration of the patch‐clamp technique to examine direct postsynaptic effects of GABAB receptor activation on SON magnocellular neurones. 2 The selective GABAB agonist baclofen reversibly inhibited voltage‐dependent Ca2+ currents elicited by voltage steps from a holding potential of −80 mV to depolarized potentials in a dose‐dependent manner. The ED50 of baclofen for inhibiting Ca2+ currents was 1.4 × 10−6 M. Baclofen did not inhibit low threshold Ca2+ currents elicited by voltage steps from −120 to −40 mV. 3 Inhibition of high threshold Ca2+ currents by baclofen was rapidly and completely reversed by the selective GABAB antagonists, CGP 35348 and CGP 55845A, when the antagonists were added at the molar ratio vs. baclofen of 10 : 1 and 0.01 : 1, respectively. It was also reversed by a prepulse to +150 mV lasting for 100 ms. 4 The inhibition of Ca2+ currents was abolished when the cells were pretreated with pertussis toxin for longer than 20 h or with N‐ethylmaleimide for 2 min. It was also abolished when GDPβS was included in the patch pipette. When GTPγS was included in the patch pipette, baclofen produced irreversible inhibition of Ca2+ currents and this inhibition was again reversed by the prepulse procedure. 5 The inhibition of N‐, P/Q‐, L‐ and R‐type Ca2+ channels by baclofen (10−5 M) was 24.1, 10.5, 3.1 and 3.6 %, respectively, of the total Ca2+ currents. Only the inhibition of N‐ and P/Q‐types was significant. 6 These results suggest that GABAB receptors exist in the postsynaptic sites of the SON magnocellular neurones and mediate selective inhibitory actions on voltage‐dependent Ca2+ channels of N‐ and P/Q‐types via pertussis toxin‐sensitive G proteins, and that such inhibitory mechanisms may play a role in the regulation of SON neurones by the GABA neurones.

Coexistence of hERG current block and disruption of protein trafficking in ketoconazole-induced long QT syndrome

Many drugs associated with acquired long QT syndrome (LQTS) directly block human ether‐a‐go‐go‐related gene (hERG) K+ channels. Recently, disrupted trafficking of the hERG channel protein was proposed as a new mechanism underlying LQTS, but whether this defect coexists with the hERG current block remains unclear. This study investigated how ketoconazole, a direct hERG current inhibitor, affects the trafficking of hERG channel protein.

PACAP increases the cytosolic Ca2+ concentration and stimulates somatodendritic vasopressin release in rat supraoptic neurons.

Pituitary adenylate cyclase activating polypeptide (PACAP)‐like immunoreactivity and its receptor mRNA have been reported in the supraoptic and the paraventricular nucleus (SON and PVN, respectively) and PACAP has been implicated in the regulation of magnocellular neurosecretory cell function. To examine the site and the mechanism of the action of PACAP in the neurosecretory cells, we measured AVP release from SON slice preparations and the cytosolic Ca2+ concentration ([Ca2+]i) from single dissociated SON neurons. PACAP at concentrations from 10−12 to 10−7 M increased [Ca2+]i in dissociated SON neurons in a dose‐dependent manner. The patterns of the PACAP‐induced [Ca2+]i increase were either sustained increase or cytosolic Ca2+ oscillations. PACAP (10−7 M) increased [Ca2+]i in 27 of 27 neurons and glutamate (10−4 M) increased [Ca2+]i in 19 of 19 SON neurons examined, whereas angiotensin II (10−7 M) increased [Ca2+]i in only 15 of 60 SON neurons examined. PACAP at lower concentrations (10−10 to 10−8 M) increased [Ca2+]i in 70–80% of neurons examined. Although the onset and recovery of the PACAP‐induced [Ca2+]i increase were slower than those observed with glutamate, the spatial distribution of the [Ca2+]i increases in response to the two ligands were similar: [Ca2+]i increase at the proximal dendrites was larger and faster and that at the center of the soma was smaller and slower. The PACAP‐induced [Ca2+]i responseswere abolished by extracellular Ca2+ removal, the l‐type Ca2+‐channel blocker, nicardipine, or by replacement of extracellular Na+ with N‐methyl d‐glucamine, and were partially inhibited by the Na+‐channel blocker, tetrodotoxin. The N‐type Ca2+‐channel blocker, ω‐conotoxin GVIA did not significantly inhibit the PACAP‐induced [Ca2+]i responses. Furthermore, PACAP (10−7 M) as well as glutamate (10−4 M) increased AVP release from SON slice preparations, and extracellular Ca2+ removal or nicardipine inhibited the AVP release in response to PACAP.

Inhibition of spontaneous inhibitory postsynaptic currents (IPSC) by noradrenaline in rat supraoptic neurons through presynaptic α2-adrenoceptors

It has been shown that noradrenergic activation has great influence on the activities of hypothalamic supraoptic neurons. No direct evidence has been reported on the presynaptic effects of adrenoceptors in the actions of noradrenaline on supraoptic neurons, although postsynaptic mechanisms have been studied extensively. In the present study, we explored presynaptic effects of noradrenaline on the supraoptic neurons by measuring spontaneous inhibitory postsynaptic currents (IPSC) with the whole-cell patch-clamp technique. Noradrenaline reduced the frequency of IPSCs in a dose-dependent (10(-9) to 10(-3) M) and reversible manner. Noradrenaline did not affect the amplitude of IPSCs at concentrations of 10(-9) to 10(-5) M, but reduced the amplitude of IPSCs at high concentrations (10(-4) and 10(-3) M). The inhibitory effects of noradrenaline were mimicked by the alpha2-agonist clonidine (10(-4) M), but not by the alpha1-agonist methoxamine (10(-4) M) nor by the beta-agonist isoproterenol (10(-4) M). Moreover, the inhibitory effects of noradrenaline on IPSCs were blocked by the non-selective alpha antagonist phentolamine (10(-4) M) or the selective alpha2-antagonist yohimbine (10(-4) M), but not by the alpha1-antagonist prazosin (10(-4) M). These results suggest that noradrena-line inhibits release of GABA from the presynaptic GABAergic terminals of the supraoptic neurons by activating presynaptic alpha2-adrenoceptors and such presynaptic mechanisms may play a role in the excitatory control of SON neurons by noradrenergic neurons.

Pre- and postsynaptic modulation of the electrical activity of rat supraoptic neurones.

The release of vasopressin and oxytocin is regulated by the electrical activity of magnocellular neurosecretory cells in the supraoptic and paraventricular nuclei, which is under the control of a great variety of neuro‐transmitters and neuromodulators. The major neural signals to the supraoptic nucleus are from excitatory glutamate inputs and inhibitory GABA inputs. In recent studies, the voltage‐clamp mode of the whole‐cell patch‐clamp technique has been applied to slice preparations from rat hypothalamus to monitor synaptic inputs to supraoptic neurones. Spontaneous excitatory and inhibitory postsynaptic currents (EPSCs and IPSCs) are abolished by CNQX and picrotoxin, respectively, but are insensitive to tetrodotoxin, indicating that they represent quantal release of glutamate and GABA, respectively, from nerve terminals of presynaptic neurones. GABA and glutamate show remarkable suppressive effects on both EPSCs and IPSCs via presynaptic GABAB and mGlu receptors, respectively. Noradrenaline, which excites supraoptic neurones via postsynaptic α1‐receptors, also suppresses IPSCs and potentiates EPSCs. On the other hand, prostaglandin E2, which excites supraoptic neurones via postsynaptic prostaglandin E2 (EP) receptors of the EP4 subclass, also suppresses IPSCs via EP3 receptors but has little effect on EPSCs. Thus pre‐ and postsynaptic mechanisms may act cooperatively to excite supraoptic neurones. Nitric oxide, which inhibits supraoptic neurones, potentiates IPSCs without affecting EPSCs. This provides another example for the preferential modulation of IPSCs of supraoptic neurones. On the other hand, PACAP, which causes a long‐lasting increase in the firing frequency via the postsynaptic receptors, has no effect on EPSCs and IPSCs, suggesting that some ligands act only at postsynaptic receptors. Thus multiple patterns for pre‐ and postsynaptic modulation are present in the supraoptic nucleus, and the electrical activity of supraoptic neurones is regulated via complex mechanisms at both pre‐ and postsynaptic sites.

Increase of urocortin-like immunoreactivity in the rat hypothalamo-neurohypophysial system after salt loading and hypophysectomy

The effect of chronic salt loading on urocortin-like immunoreactivity (Ucn-IR) was investigated in the rat hypothalamo-neurohypophysial system. In control rats a few Ucn-IR neurons were observed scattered throughout the supraoptic nucleus (SON) and few in the paraventricular nucleus (PVN). A small number of Ucn-IR fibers were observed scattered in the median eminence (ME) and the posterior pituitary. However, after 5 days of chronic administration of 2% saline, a marked increase in the number of Ucn-IR perikarya and fibers was observed in the PVN and the SON. Additionally, Ucn-IR varicosities and fibers were found in the internal zone of the ME and in the posterior pituitary. To confirm the findings and examine the possible involvement of anterior pituitary function in synthesis of Ucn, surgical hypophysectomized rats were used. Five days after hypophysectomy, a marked increase in Ucn-IR was observed in the PVN, the SON, and both the internal and the external zone of the ME. These results suggest that Ucn in the hypothalamo-neurohypophysial system may be involved in the regulation of salt balance, and possibly in the stimulation of ACTH release from the anterior pituitary.

The Mechanism of Inhibitory Actions of Propofol on Rat Supraoptic Neurons

BACKGROUND In the perioperative period, plasma osmotic pressure, systemic blood pressure, and blood volume often change dramatically. Arginine vasopressin is a key factor in the regulation of these parameters. This study was performed to evaluate the direct and the mechanism of the actions of propofol on arginine vasopressin release from magnocellular neurosecretory neurons in the rat supraoptic nucleus. METHODS Somatodendritic arginine vasopressin release from supraoptic nucleus slice preparations was measured by radioimmunoassay. Ionic currents were measured using the whole-cell mode of the patch-clamp technique in supraoptic nucleus slice preparations or in single dissociated supraoptic nucleus neurons of the rat. RESULTS Propofol at concentrations greater than 10(-5) M inhibited the arginine vasopressin release stimulated by potassium chloride (50 mM). This inhibition by propofol was not reversed by picrotoxin, a gamma-aminobutyric acid(A)(GABA(A)) receptor antagonist, whereas arginine vasopressin release induced by glutamate (10(-3) M) was also inhibited by propofol at a clinically relevant concentration (10(-6) M). The latter effect was reversed by picrotoxin. Propofol evoked Cl- currents at concentrations ranging 10(-6) to 10(-4) M. Propofol (10(-6) M) enhanced the GABA (10(-6) M)-induced current synergistically. Moreover, propofol (10(-6) M) prolonged the time constant of spontaneous GABA-mediated inhibitory postsynaptic currents. Furthermore, propofol (10(-5) M and 10(-4) M) reversibly inhibited voltage-gated Ca2+ currents, whereas it did not affect currents induced by glutamate (10(-3) M). CONCLUSIONS Propofol inhibits somatodendritic arginine vasopressin release from the supraoptic nucleus, and the enhancement of GABAergic inhibitory synaptic inputs and the inhibition of voltage-gated Ca2+ entry are involved in the inhibition of arginine vasopressin release.

Ionotropic and Metabotropic Glutamate Receptor Agonist-Induced [Ca2+]i Increase in Isolated Rat Supraoptic Neurons

In the present study, the effects of glutamate and of agonists for ionotropic and metabotropic glutamate receptors on intracellular Ca2+ concentration ([Ca2+]i) were investigated in neurons of the rat supraoptic nucleus (SON). We used the intracellular Ca2+ imaging technique with fura‐2, in single magnocellular neurons dissociated from the SON of rats. Glutamate (10−6−10−4 M) evoked a dose‐dependent increase in [Ca2+]i. The glutamate agonists exerted similar effects, although with some differences in the characteristics of their responses. The [Ca2+]i response to NMDA was smaller than those of glutamate or the non‐NMDA receptor agonists, AMPA and kainate, but was significantly enhanced by the removal of extracellular Mg2+. Glutamate, as well as quisqualate, an agonist for both ionotropic and metabotropic glutamate receptors, evoked a [Ca2+]i increase in a Ca2+‐free condition, suggesting Ca2+ release from intracellular Ca2+ stores. This was further evidenced by [Ca2+]i increases in response to a more selective metabotropic glutamate receptor agonist, t‐ACPD, in the absence of extracellular Ca2+. Furthermore, the quisqualate‐induced Ca2+ release was abolished by the selective metabotropic glutamate receptor antagonist, (S)‐4‐carboxyphenylglycine. The results suggest that metabotropic glutamate receptors as well as non‐NMDA and NMDA receptors are present in the SON neurons, and that activation of the first leads to Ca2+ release from intracellular Ca2+ stores and the activation of the latter two types induces Ca2+ entry. These dual mechanisms of Ca2+ signalling may play a role in the regulation of SON neurosecretory cells by glutamate.

Upregulation of the expression of vasopressin gene in the paraventricular and supraoptic nuclei of the lithium-induced diabetes insipidus rat.

The expression of arginine vasopressin (AVP) gene in the paraventricular (PVN) and supraoptic nuclei (SON) was investigated in rats with lithium (Li)-induced polyuria, using in situ hybridization histochemistry and radioimmunoassay. The male Wistar rats consuming a diet that contained LiCl (60 mmol/kg) for 4 weeks developed marked polyuria. The Li-treated rats produced a large volume of hypotonic urine with low ionic concentrations. Plasma sodium concentrations were found to be slightly increased in the Li-treated rats compared with those in controls. Plasma concentration of AVP and transcripts of AVP gene in the PVN and SON were significantly increased in the Li-treated rats compared with controls. These results suggest that dehydration and/or the activation of visceral afferent inputs may contribute to the elevation of plasma AVP and the upregulation of AVP gene expression in the PVN and the SON of the Li-induced diabetes insipidus rat.