Rebecca J. Naukam

Effects of calcium channel blocking agents on calcium and centrilobular necrosis in the liver of rats treated with hepatotoxic agents

Carbon tetrachloride, chloroform, dimethylnitrosamine, thioacetamide or acetaminophen was each administered to rats in a single hepatotoxic dose. Nifedipine, verapamil or chlorpromazine was administered in association with the hepatotoxic agents to determine if calcium channel blocking agents would prevent an increase in liver cell calcium associated with hepatotoxicity and to determine if these agents would protect against the development of centrilobular necrosis. Following a latent period different for each toxic agent, a 4- to 18-fold increase in liver cell calcium content had occurred by 24 hr. The calcium increase and the centrilobular necrosis (mean histologic score) were correlated. A relatively high calcium to necrosis ratio was obtained with dimethylnitrosamine, thioacetamide and acetaminophen. A lesser calcium to necrosis ratio was obtained with chloroform and carbon tetrachloride, the two toxic agents that destroyed the intracellular calcium sequestration activity of the liver endoplasmic reticulum. Nifedipine or chlorpromazine, administered prior to and 7 hr after the toxic agent, completely prevented the centrilobular necrosis caused by thioacetamide, carbon tetrachloride and acetaminophen; almost completely prevented necrosis with dimethylnitrosamine; and provided partial protection against chloroform toxicity. Two doses of verapamil provided partial protection against necrosis when carbon tetrachloride was the toxic agent and provided almost complete protection with dimethylnitrosamine. A reduction in liver cell calcium was associated with the protective action of the three calcium channel blocking agents. These findings are compared with earlier studies of the protective effects of calcium channel blocking agents in cardiac ischemia.

Immunoreactive substance P is decreased in saliva of patients with chronic back pain syndromes

Substance P, a neuropeptide associated with pain perception, is widely distributed in the central nervous system and is decreased in the cerebrospinal fluid of chronic pain patients as compared with that of healthy human volunteers. In this study, we have demonstrated the presence of immunoreactive substance P in saliva and further, that both saliva and plasma levels of immunoreactive substance P are lower in patients with chronic low back pain than in healthy human volunteers. To our knowledge, this is the first time that substance P has been identified in human saliva. These findings, together with the noninvasive nature of saliva collection, suggest that substance P in saliva may be useful as an alternative neurochemical correlate of chronic low back pain when collection of cerebrospinal fluid and plasma samples for substance P analysis is unacceptable or inappropriate.

Effects of calcium channel blocking agents on membrane microviscosity and calcium in the liver of the carbon tetrachloride treated rat

Membrane microviscosity was determined from the polarized fluorescence of diphenylhexatriene in plasma membranes and microsomes prepared from the liver of carbon tetrachloride treated rats. It was greatly depressed between 12 and 24 hr after the administration of the carbon tetrachloride. Depression of microviscosity was also seen in the liposomes which were prepared from these membranes. There were decreases in phospholipid content and phospholipid methyltransferase activity, but these changes did not appear to explain the decreased microviscosity. A large accumulation of calcium occurred in the liver cells between 12 and 24 hr after the administration of carbon tetrachloride. Chlorpromazine, verapamil and nifedipine, when administered prior to the carbon tetrachloride, partially reduced the later accumulation of calcium and reduced the degree of histological damage observed. When these agents were administered 12 hr after the administration of carbon tetrachloride, they did not reduce the subsequent accumulation of calcium. When administered prior to and 7 hr after carbon tetrachloride, they had a small but potentially significant effect on the microviscosity change. It is suggested that at low levels of microviscosity a critical threshold may exist below which entry of calcium into the cell is poorly controlled and that calcium channel blocking agents may be ineffective if administered at a time when membrane microviscosity is very low. Tissue calcium accumulation was associated with visible cell damage.

The inhibitory effect of metoclopramide on plasma cholinesterase activity

The in vitro effect of metoclopramide on plasma cholinesterase (PCHE) activity was studied to investigate a mechamistn for metoclopramide-induced prolongation of succinylcholine action. The mean PCHE of the control samples was 0.86 ° 0.02 unit·ml-1. PCHE activity in the presence of metoclopramide, at concentrations of 0.05, 0.10, 0.50,1.0, 2.5 and 5.0 μg·ml-1, was reduced to 0.78 ± 0.02, 0.69 ± 0.04,0.50 ± 0.03,0.39 ± 0.02, 0.24 ± 0.01 and 0.15 ± 0.01 unit ·ml-1, respectively. Our data demonstrated that PCHE activity was significantly depressed by metoclopramide at all concentrations studied (p < 0.001). Our data also show that the concentration of metoclopramide required to inhibit 50 per cent of PCHE activity (I50) was 0.8 μg·ml-1 (2.4 × 10-6 M). We recommend caution when succinylcholine and or ester type local anaesthetics are administered to patients who are also receiving metoclopramide, especially in high doses.RésuméĽeffet in vitro de la métoclopramide sur ľactivité de la cholinestérase plasmatique (PCHE) a été étudié afin ďinvestiguer le mécanisme de la prolongation de ľaction de la succinylcholine induite par la métoclopramide. La valeur moyenne des échantillons de contrôle de PCHE était de 0.86 ± 0.02 unités · ml-1. Ľactivité du PCHE en présence de métoclopramide à des concentrations de 0.05, 0.10, 0.50,1.0. 2.5 et 5.0 μg·ml-1, a été réduiteà0.78 ± 0.02, 0.69 ± 0.04, 0.50 ± 0.03, 0.39 ± 0.02, 0.24 ± 0.01 et 0.15 ± 0.01 unités·ml-1 respectivement. Nos données ont démontré que ľactivité de la PCHE était significativement diminuée par la métoclopramide à toutes les concentrations étudiées (p < 0.001). Nos données ont aussi démontré que la concentration de métoclopramide requise afin ďinhiber 50 pour cent de ľactivité de la PCHE (150) était de 0.8 μg · ml-1 (2.4 × 10-6 M). On recommande la précaution quand la succinylcholine ou anesthésique local type ester est administré aux patients qui reçoivent aussi la métoclopramide, spécialement à hautes doses.

The effect of procainamide on plasma cholinesterase activity

The in vitro effect of procainamide on plasma cholinesterase (PCHE) activity in the plasma often normal ASA physical status I patients was studied using a kinetic method. The mean plasma cholinesterase activity without procainamide (control) was 0.90 ± 0.09 units.ml-1 The dibucaine numbers of all the samples were in the normal range of 78 to 86, indicating normal genotypes. The mean plasma cholinesterase activity, in the presence of procainamide in concentrations of 5.0, 10.0, 20.0 and 40.0 µ.ml-1, was reduced to 0.73 ± 0.04. 0.61 ± 0.03, 0.45 ± 0.02, and 0.36 ± 0.01 units.mt-1, respectively. At therapeutic concentrations of 4 to 12 µg.ml-1, procainamide inhibited cholinesterase activity 15 to 30 per cent. The authors also showed that the concentration of procainamide required to inhibit 50 per cent of plasma cholinesterase activity was 20 µg.mt-1 (Iso). The authors conclude that procainamide when tested in vitro had a statistically significant depressant effect on plasma cholinesterase activity at all the concentrations studied.RésuméOn a étudié ľeffet in vitro de la procaïnamide sur ľactivité de la cholinestérase plasmatique (PCHE) dans le plasma de dix patients normaux de statut physique ASA I, à ľaide ďune méthode cynétique. Ľactivité de la cholinestérase plasmatique moyenne sans procainamide (groupe témoin) était de 0.90 ± 0.09 unités.ml-1. Les nombres de dibucaine, pour tous les échantillons, se situaient dans ľéchelle normale de 78 à 86 dénotant, de ce fait, des génotypes normaux. La présence de procainamide dans des concentrations de 5.0, 10.0, 20.0 et 40.0 µg.ml-1, réduisait ľactivité de la cholinestérase plasmatique moyenne à 0.73 ± 0.04, 0.61 ± 0.03, 0.45 ± 0.02, et 0.36 ± 0.01 unités.ml-1, respectivement. Des concentrations thérapeutiques de 4 à 12 µg.ml-1 de procaïnamide inhibaient ľactivité de la cholinestérase de 15 à 30 pour cent. Les auteurs ont aussi démontré que la concentration de procaïnamide requise pour inhiber 50 pour cent de ľactivité de la cholinestérase plasmatique était de 20 µg.ml-1 (I50). Les auteurs concluent que la procaïnamide testée in vitro a un effet dépressif statistiquement significatif sur ľactivité de la cholinestérase plasmatique à toutes les concentrations étudiées.

In vitro effects of fluoride and bromide on pseudocholinesterase and acetylcholinesterase activities

The in vitroeffects of two metabolites ofinhalational anaesthetics, fluoride and bromide, on pseudocholinesterase (PCHE) and acetylcholinesterase (ACHE) activities in the blood samples of seven healthy patients were studied. The PCHE and ACHE activities were determined by kinetic spectrophotometric methods. Fluoride at the levels achieved with clinical concentrations of enjlurane and sevoflurane (25–75 μM · L−1) inhibited PCHE activity by 28–65 per cent (P < 0.01) and ACHE activity by less than five per cent (P > 0.05). Bromide at the levels achieved with clinical concentrations ofinhalational anaesthetics had no significant effect on either PCHE or ACHE activity. We recommend caution when succinylcholine and/or ester type local anaesthetics are used in the immediate postoperative period following enflurane or sevoflurane anaesthesia. We also recommend that blood drawing for PCHE activity be delayed at least until 24 hr following enflurane or sevoflurane anaesthesia.RésuméLes effets in vitrode deux métabolites anesthésiques d’inhalation, le fluorure el le bromure sur les activités de la pseudocho-linesterase (PCHE) el l’acetylcholinesterase (ACHE) out été étudié sur les échantillons sanguins de sept patients en bonne sante. Les activités de la PCHE et l’ACHE out été déterminées par spectrophotométrie. Les niveaitx de fluorure atteints avec des concentrations d’enjlurane et de sevoflurane à des concen-trations utilisees en clinique (25–75 μM · L−1) out inhibé l’activite de la PCHE de 28–65 pour cent (P < 0.0l) et l’activité de l’ACHE de moins de cinq pour cent (P > 0.05). Les niveaux de bromure atteints aux concentrations anesthesiques utilisees en clinique n’ont eu aucun effet significatif sur les activites de la PCHE ou de l’ACHE. On recommande la précaution lors de l’utilisation de la succinylcholine etlou les anesthésiques locaux dans la période post-opératoire immédiate après l’anesthésique à l’enflurane ou le sevoflurane. On recommande aussi que la prise d’échantillon sanguin pour l’activite de la PCHE soit retardée d’au moins 24 heures après l’anesthésie à l’enflurane ou la sevoflurane.

The effects of neurokinin A, neurokinin B, and eledoisin on substance P analysis

Commercial sources for neuropeptide radioimmunoassays have made this sensitive tool available to clinical investigators for monitoring the potential involvement of neuropeptides in pain modulation. We measured substance P-like immunoreactivity in the plasma, saliva, and pericardial fluid of subjects with and without pain (chronic and acute) to determine if substance P levels are altered. Some recent studies have suggested that substance P in various body fluids may be a correlate of chronic pain. To test this correlation it is important to ensure that the assay is measuring what it was designed to measure. Therefore, the influence of three tachykinins on the analysis of substance P concentrations was assessed with a commercially available radioimmunoassay kit. A small (approximately 2 to 6%), apparently nonspecific elevation in measured substance P was found when alpha-neurokinin, beta-neurokinin, or eledoisin was incubated with substance P and its antibody. Our results also indicate an apparent specific affinity of the substance P antibody for alpha-neurokinin (above 1,000 pg/ml) and beta-neurokinin (above 5,000 pg/ml). Substance P levels in the body fluids we tested ranged from 0.47 to 62.88 pg/mg protein (47.4 to 230.8 pg/ml). Levels of the tested tachykinins have not been determined in body fluids. If alpha-neurokinin or beta-neurokinin is found to be present in high concentrations in these fluids, this commercially available substance P kit may overestimate substance P levels. The concentrations of tachykinins necessary to interfere specifically with the assay are 10- to 100-fold higher than substance P in body fluids. If the other tachykinins are present at concentrations similar to the substance P levels, we would not expect them to interfere substantially with the accuracy of our measurements of substance P using this assay.

Maternal tobacco smoking and changes in amino acid uptake by human placental villi: Induction of uptake systems, gammaglutamyltranspeptidase and membrane fluidity

Maternal smoking depressed the active uptake of amino acids by human placentae and lowered their levels in the placenta and umbilical vein. During starvation of cells for amino acids, more amino acid carriers are induced and incorporated into the plasma membrane. A question arises whether there could be similar changes due to maternal smoking in the placental amino acid uptake carrier systems. Therefore, the characteristics of (a) the uptake of 2-amino[I-14C]-isobutyric acid (AIB) by isolated placental villi, (b) gammaglutamyltranspeptidase (GGTP), a critical enzyme of the gammaglutamyl cycle (GGC) for the uptake of amino acids in human placenta, and (c) lipid structural parameters (reciprocal of fluidity), by steady state fluorescence polarization of plasma membrane vesicles of microvilli (MV) and microsomal membranes (MM) of umbilical and chorionic plate arteries of placentae of smoking and non-smoking mothers were investigated. The above investigations gave the following results: (a) Washed placental villi of smokers exhibited higher capacity for AIB uptake than those of non-smokers. The higher uptake capacity was mainly due to increase in Vmax for AIB uptake in smokers. Km increased for placental AIB uptake in smokers. (b) Maternal smoking lowered GGTP activity of MV by decreasing its Vmax. Therefore, maternal smoking decreases the formation of gammaglutamyl-amino acid (GGAA) on the surface of trophoblast which are absorbed by the trophoblast. The degree of absorption of GGAA is considered as an inverse environmental signal for the cell to regulate amino acid transport systems. Maternal smoking seems to decrease the formation and absorption of GGAA and thereby induce the formation of new carriers for AIB uptake. (c) Maternal smoking increased the values for lipid structural order parameters and microviscosity of MV and induced tolerance against fluidization by ethyl alcohol in MM of umbilical and chorionic arteries. The alterations could increase Km for AIB uptake system and decrease the sensitivity of umbilical and chorionic arteries to vasoconstrictive substances like 5-hydroxytryptamine and catecholamine which are released by nicotine. All these changes tend to overcome the deficits produced in placental amino acid transport and satisfy the demands of the growing fetus for amino acids.

In Vivo Effects of Nitrendipine on Hepatotoxic Cell Injury in Rats

Carbon tetrachloride, thioacetamide, and other hepatotoxic agents were each administered to rats in a single toxic dose. Experimental studies were designed to test the following hypothesis. The cell injury caused by some hepatotoxic agents is associated with the entry of calcium into the liver cells. Some calcium channel blocking agents, such as nitrendipine, will reduce both the entry of calcium and the associated liver-cell injury. In the following studies the hepatotoxic agents caused a rise in liver-cell calcium content that was associated in its time course with the development of contrilobular necrosis. Nitrendipine, and several other calcium channel blocking agents that were tested, reduced both the calcium increase and the centrilobular necrosis. The in-vivo experimental evidence suggests that this calcium increase is not solely the consequence of a calcium influx that follows cell necrosis. The ratio of the increase in calcium to-cell necrosis varied with the specific group of hepatotoxic agents employed. With some toxic compounds the calcium channel blocking agents diminished the calcium increase, but virtually eliminated the cell necrosis. The corollary is that a part of the calcium increase precedes the necrosis and may play a role in the development of cell injury. Acute and chronic ingestion of ethanol was not associated with an increase in liver-cell calcium content. Possible mechanisms and potential limitations of the observed protection from cell injury are discussed. These findings support the existing evidence that calcium channel blocking agents employed in treatment of cardiovascular disease may reduce the associated cell injury.