Antinociceptive Effects of Amiloride and Benzamil in Neuropathic Pain Model Rats

Amiloride and benzamil showed antinocicepitve effects in several pain models through the inhibition of acid sensing ion channels (ASICs). However, their role in neuropathic pain has not been investigated. In this study, we investigated the effect of the intrathecal amiloride and benzamil in neuropathic pain model, and also examined the role of ASICs on modulation of neuropathic pain. Neuropathic pain was induced by L4-5 spinal nerve ligation in male Sprague-Dawley rats weighing 100-120 g, and intrathecal catheterization was performed for drug administration. The effects of amiloride and benzamil were measured by the paw-withdrawal threshold to a mechanical stimulus using the up and down method. The expression of ASICs in the spinal cord dorsal horn was also analyzed by RT-PCR. Intrathecal amiloride and benzamil significantly increased the paw withdrawal threshold in spinal nerve-ligated rats (87%+/-12% and 76%+/-14%, P=0.007 and 0.012 vs vehicle, respectively). Spinal nerve ligation increased the expression of ASIC3 in the spinal cord dorsal horn (P=0.01), and this increase was inhibited by both amiloride and benzamil (P<0.001 in both). In conclusion, intrathecal amiloride and benzamil display antinociceptive effects in the rat spinal nerve ligation model suggesting they may present an alternative pharmacological tool in the management of neuropathic pain at the spinal level.

Effect of aldosterone on the amplification of oncolytic vaccinia virus in human cancer lines / 대한간학회지

BACKGROUND/AIMS: JX-594 is an oncolytic virus derived from the Wyeth vaccinia strain that causes replication-dependent cytolysis and antitumor immunity. Starting with a cross-examination of clinical-trial samples from advanced hepatocellular carcinoma patients having high levels of aldosterone and virus amplification in JX-594 treatment, we investigated the association between virus amplification and aldosterone in human cancer cell lines. METHODS: Cell proliferation was determined by a cell-counting-kit-based colorimetric assay, and vaccinia virus quantitation was performed by quantitative polymerase chain reaction (qPCR) and a viral plaque assay. Also, the intracellular pH was measured using a pH-sensitive dye. RESULTS: Simultaneous treatment with JX-594 and aldosterone significantly increased viral replication in A2780, PC-3, and HepG2 cell lines, but not in U2OS cell lines. Furthermore, the aldosterone treatment time altered the JX-594 replication according to the cell line. The JX-594 replication peaked after 48 and 24 hours of treatment in PC-3 and HepG2 cells, respectively. qPCR showed that JX-594 entry across the plasma membrane was increased, however, the changes are not significant by the treatment. This was inhibited by treatment with spironolactone (an aldosterone-receptor inhibitor). JX-594 entry was significantly decreased by treatment with EIPA [5-(N-ethyl-N-isopropyl)amiloride; a Na+/H+-exchange inhibitor], but aldosterone significantly restored JX-594 entry even in the presence of EIPA. Intracellular alkalization was observed after aldosterone treatment but was acidified by EIPA treatment. CONCLUSIONS: Aldosterone stimulates JX-594 amplification via increased virus entry by affecting the H+ gradient.

Myocardial protection by Ethyl-isopropyl amiloride, a specific NA[+] -H[+] exchange inhibitor, following hemorrhagic shock

Hemorrhagic shock and resuscitation is well known to result in myocardial dysfunction and injury. Stimulation of the Na[+] -H[+] exchanger plays an important role in the pathway of myocardial injury. The purpose of the present study was to examine the protective effects of blocking the cardiac Na[+] -H[+] exchange, using 100mM ethyl-isopropyl amiloride [EIPA], a specific Na[+] -H[+] exchanger blocker, on myocardial contractile function on ex vivo resuscitation of isolated rat heart following one hour of hemorrhagic shock. Sprague- Dawley rats were assigned to hemorrhage, hemorrhage + EIPA, sham hemorrhage and sham hemorrhage + EIPA groups. Rats were hemorrhaged for one hour. Hearts were harvested and ex vivo treated and resuscitated by perfused in the Langendorff System. Myocardial function was determined. The results showed that inhibition of the Na[+] -H[+] exchanger using EIPA improved the post-resuscitation myocardial contractile function. Blocking the Na[+] -H[+] exchanger using 100mM EIPA following 60 minutes of hemorrhagic shock improved myocardial function

Potentiation of spermicidal activity of 2',4'-dichlorobenzamil by lidocaine.

The present investigation was designed to study the spermicidal activity of lidocaine, a membrane stabilizer, and its combination with 2',4'-dichlorobenzamil hydrochloride, a Na+-Ca2+ exchange inhibitor, on human semen and spermatozoa separated from semen. Both drugs per se produced dose- and time-dependent reduction in motility of ejaculated human sperm. Lidocaine was found to potentiate the spermicidal activity of benzamil resulting in significant decrease in time for producing complete loss of ejaculated sperm motility. Sperm revival test revealed irreversible loss of sperm viability indicating a spermicidal rather than spermiostatic action by both the drugs. Furthermore, both benzamil (10-40 mM) per se and benzamil-lidocaine combination (0.5 and 16 mM) produced contraception in rabbit model.

Efecto de la amilorida y su derivado diclorobenzamil sobre la auricula aislada del acure (Cavia porcellus): interaccion con otras maniobras inotropicas / Effect of amiloride and dichlorobenzamil derivative in the guinea pig atria: interaction with other inotropic mechanisms

Se estudiaron los efectos cronotrópico e inotrópico de la Amilorida (AMI) y la dicloro-benzamil-Amilorida (DCB-AMI) sobre las aurículas aislada del acure, así como la interacción de estas drogas con la beta-metil-digoxina (BM_DIGO), la epinefrina y la disminución del potasio extracelular (de 4 a 1 mM). La AMI (1 mM) causa un efecto inotrópico positivo y cronotrópico negativo, independientes del sistema autonómico. La DCB-AMI causa um efecto bimodal sobre la fuerza de contracción: la aumenta a bajas dosis pero la disminuye a concentraciones mayores de 10(-6) M. También disminuye levemente la frecuencia sinusal. El efecto de la AMI sobre el automatismo sinusal no es alterado por la BM-DIGO. En cambio, la AMI ((10(-3 M) disminuye el efecto inotrópico positivo de la BM-DIGO e incrementa la dosis tóxica en preparaciones aisladas. La curva dosis-respuesta a la epinefrina no varía en presencia de AMI. Resultados similares se obtuvieron con DCB-AMI (2 x 10(-7 M). El incremento de contractilidad que se observa al disminuir la concentración extracelular de potasio a 1 mM no se altera en presencia de AMI. La actividad de la Na+/K+ ATPasa dependiente de Mg++ de la fracción microsomal obtenida del corazón del acure disminuye en 10 por ciento aproximadamente en presencia de AMI (1nM). Por otra parte, el efecto inhibitorio sobre la enzima obtenido con ouabaína no varía con esta droga. En conclusión, nuestros resultados sugieren múltiples efectos de la AMI y DCB-AMI sobre el corazón del acure. La inhibición del intercambiador Na+/Ca++ explica solo parte de ellos; el bloqueo de los canales lentos parece fundamental para explicar nuestras observaciones.

Effects of dichlorobenzamil, a sodium-calcium exchange inhibitor, on the calcium paradox and the sodium withdrawal contractures of frog atrial muscle

The effects of dichlorobenzamil (DCB), an amiloride derivative and potent inhibitor of Na-Ca exchange in cardiac sarcolemmal vesicles and isolated cardiac myocytes, were investigated in two paradigms involving Na-Ca exchange, namely the Ca2+ paradox and the Na + - withdrawal contractures of frog atrial muscle strips. Pretreatment with DCB (10-100 micronM) inhibited in a dose-dependent manner the contractures elicited by reexposure of the atrial strips to the control Ringer solution after a 5-20 min equilibration with a Ca2 + - free saline (Ca2 + - readmission contractures; Ca2 + paradox). These contracture were not inhibited, however, when DCB was applied after the preparation had been exposed to the Ca2 + - free saline, but before the reexposure to the control Ringer solution. DCB (10-100 micronM) did not inhibit the contractures elicited by Na + - deficient saline (Na + - withdrawal contractures) in atrial strips pretreated or not with acetylstrophantydin. This result suggests that, under our experimental conditions, DCB falied to substantially inhibit the Ca2 + influx mediated by Na-Ca exchange. The duration of the plateu of the action potentials of atrial cells equilibrated with Ca2 + - free saline was reduced from 1,42 ñ 0,27 s to 0,61 ñ 0,13 s by 50 micronM DCB (P<0.001). This was atributed to blockade of Na + currents through modified L-type Ca2 + channels. It is proposed that shortening of the Na + - dependent action potentials can account for the inhibition of the Ca2 + - readmission contractures, because these contractures have a steep dependence on the Na + influx and intracellular Na + accumulation that occurs during the Ca2 + - free period. The results of this study support the conclusion thatDCB has multiple effects on heart muscle, including a potent blockade of Ca2 + channels, and its use as a selective inhibitor of Na-Ca exchange in cellular systems in un unwarranted