Cholinergic mechanisms of analgesia produced by physostigmine, morphine and cold water swimming.

This study concerns the cholinergic involvement in three experimental procedures which produce analgesia. Rats were given one of seven treatments: saline (1.0 ml/kg, i.p.); morphine sulfate (3.5, 6.0 or 9.0 mg/kg, i.p.); physostigmine salicylate (0.65 mg/kg, i.p.); warm water swim (3.5 min at 28 degrees C); and cold water swim (3.5 min at 2 degrees C). Each rat was tested on a hot plate (59.1 degrees C) once prior to and 30 min after treatment. Immediately after the last test the rats were killed with focussed microwave radiation. Levels of acetylcholine (ACh) and choline (Ch) in six brain areas (brain stem, cerebral cortex, hippocampus, midbrain, cerebellum and striatum) were analyzed by gas chromatograph-mass spectrometer. Morphine (9.0 mg/kg), physostigmine and cold water swimming caused significant analgesia. Morphine elevated the levels of ACh in the cerebellum and striatum, cold water swimming--in the cerebellum, striatum and cortex, and physostigmine--in the striatum and hippocampus. Levels of choline were elevated by morphine in the cerebellum, cortex and hippocampus, while cold water swimming elevated levels of choline in the cerebellum, cortex, striatum and hippocampus. Physostigmine did not change levels of choline in any of the brain areas studied. These data suggest that the analgetic effects of morphine or cold water swimming may be mediated by components of the cholinergic system that differ from those involved in the analgetic effects of physostigmine.

The effect of adrenalectomy and dexamethasone on the antinociceptive effects of physostigmine.

The tail-flick procedure was used to study the antinociceptive effects of physostigmine in adrenalectomized and sham-operated rats. At 5 days after surgery, they were tested 30 min after either 0.32 or 0.45 mg/kg IP physostigmine. Adrenalectomized animals showed significantly greater elevation of TF scores from predrug latencies than the sham controls at both doses of physostigmine. Following 3 days of dexamethasone replacement therapy on days 18, 19, and 20 post-surgery the antinociceptive effects of physostigmine were uniformly attenuated across doses or surgical groups. On the other hand, animals receiving saline injection instead of dexamethasone did not manifest any reduction of the physostigmine antinociceptive effect. The potentiation by adrenalectomy and the reduction following dexamethasone of the antinociceptive effects of physostigmine suggest that these effects may be mediated through hypothalamic-pituitary-adrenal mechanisms and are consistent with beta-endorphin-induced sensitization of opiate or cholinergic receptors.

Stereoacuity development in children with normal binocular single vision.

We made 344 stereoacuity determinations (Titmus Stereotest) on 321 children, ages 1 1/2 to 13 years, who had normal binocular single vision tested by other factors. The data showed a gradual improvement in stereoacuity scores with increasing age--up to age 9--when a normal stereoacuity of 40 seconds of arc was consistently found. The lower limits of stereoacuity compatible with normal binocular single vision were 3 1/2 years, 3,000 seconds; 5 years, 140 seconds: 5 1/2 years, 100 seconds; 6 years, 80 seconds; 7 years, 60 seconds; and 9 years, 40 seconds.

Age-related differences in soman toxicity and in blood and brain regional cholinesterase activity.

The toxicity (lethality, acute toxic signs and body weight loss) of the irreversible ChE inhibitor soman was assessed in four groups of male rats differing in age: 30, 60, 120 and 240 days old. Plasma and brain regional ChE activity profiles were also studied in these groups. All measures of the toxicity of soman were found to increase with age. The calculated 24-hr LD50s were 110, 87, 66 and 59 micrograms/kg, IM, for 30-, 60-, 120- and 240-day-old rats, respectively. A significant and positive age-related effect on toxic sign rating scores was observed at one hr following soman injection. Furthermore, during a 14-day postsoman observation period, it was observed that young rats had less initial weight loss and more rapid, sustained recovery of growth than older animals. Survivors from the two oldest age groups did not recover to baseline body weights by the end of the 14-day observation period. Basal level of plasma ChE activity did not change significantly with age, while brain regional ChE showed two distinct age-dependent patterns: a linear decrease in the brainstem, midbrain and cerebellum and an inverted U-shaped change in the cortex, hippocampus and striatum. Our data suggest a relationship between soman toxicity and the aging process, but fails to demonstrate a definite relationship between soman toxicity and basal ChE activity in blood and brain of rats.

Conditioned taste aversion and cholinergic drugs: pharmacological antagonism.

The effectiveness of drugs as unconditioned stimuli (UCSs) in the conditioned taste aversion (CTA) procedure may be influenced by specific pharmacological antagonism. The present studies examined the UCS effects of two carbamates, physostigmine salicylate (PS) and pyridostigmine bromide (PB), and three anticholinergic compounds, atropine methyl nitrate (AMN), atropine sulfate (AS), and benactyzine hydrochloride (BH). Individual drugs, as well as combinations of the carbamates and the anticholinergics, were studied in a two-bottle procedure in rats. The lowest effective doses for eliciting significant CTAs were PS, 0.32 mg/kg; PB, 1.00 mg/kg; AMN, 0.04 mg/kg; AS, 0.07 mg/kg and BH, 0.90 mg/kg, IP. Combinations of PS with either AMN or BH were mutually antagonistic as UCSs, whereas PS with AS was not. PB with AMN, but not with AS, also showed antagonism in the procedure. The present results suggest that the CTA procedure is well-suited for direct examination of cholinergic drug effects and may also be used to explore interactions of different classes of cholinergic drugs.

The effects of choline on soman-induced analgesia and toxicity.

The effects of acute choline (Ch) administration on the hot plate (HP) analgesia produced by soman and by morphine and on the toxicity produced by soman were examined in male rats. Morphine (9.0 mg/kg, IP) and soman (80 micrograms/kg, SC), but not Ch (60 mg/kg, IP), produced HP analgesia. Pretreatment with Ch (100 mg/kg, IP; given 20 min earlier) reduced morphine and soman analgesia by 68% and 38% respectively. Ch (LD50 = 405.8 mg/kg, IP) at doses above 200 mg/kg rapidly produced signs of cholinergic stimulation which disappeared by one hr. A fixed dose of Ch (100 mg/kg) altered neither the expression of toxic cholinergic signs produced by varying doses (from 75.9 to 151.4 micrograms/kg) of soman, nor the 24-hr LD50 (124.0 micrograms/kg, SC) of soman. Doses of Ch ranging from 213.8 to 269.2 mg/kg when given 40 min prior to a fixed dose (93.3 micrograms/kg) of soman resulted in persistent signs of cholinergic hyperstimulation, but again, did not affect the mortality produced by soman. The results demonstrate that acute Ch pretreatment increased the severity of cholinergic stimulation and reduced the HP analgesia, but were not accompanied by potentiation or antagonism of the lethal action of soman.

A rapid in vitro assay of cellular chemomigration in an epithelial carcinoma cell line.

We have studied the chemomigration activity of an epithelial carcinoma cell line using a modified 96-well Boyden chamber apparatus consisting of upper and lower wells separated by an 8-microns pore polycarbonate filter. Cells from the malignant squamous carcinoma cell line A-431 were plated in the upper wells over a collagen IV-coated filter. In chemokinesis assays, the cells were allowed to migrate toward NIH 3T3 fibroblast-conditioned medium or control media in the lower wells for 6 hours at 37 degrees C with 10% CO2. A-431 cells preferentially migrate across the barrier toward conditioned media but not control media. Control normal keratinocytes showed no migration. A highly metastatic melanoma cell line and poorly metastatic melanoma cell line, in which chemomigration has been shown previously to correlate with metastatic potential, were used as positive and negative cellular controls. This system provides a rapidly quantifiable method by which the invasion characteristics of multiple cell lines can be studied simultaneously in a single assay using the 96-well format.

Comparison of hematologic consequences and efficacy of p-aminophenones in mice.

Controlled methemoglobin (MHb) formation is one strategy employed to counter cyanide (CN) toxicity. Currently available MHb formers present certain drawbacks and limitations. The purpose of this study was to characterize, in mice, the hematologic effects of the MHb-forming compound p-aminopropiophenone (PAPP), and two structurally-related p-aminophenones, p-aminoheptanoylphenone (PAHP) and p-aminooctanoylphenone (PAOP). Although these three p-aminophenones have been shown previously to be efficacious as pretreatments against CN, a more complete understanding of their hematologic effects is lacking. In addition, because the active form of PAPP has been shown to be its N-hydroxy metabolite, the N-hydroxy metabolites of PAPP, PAHP and PAOP were also tested. Using a hemoximeter, blood samples obtained -2 to +180 min relative to intramuscular (i.m.) or intraperitoneal (i.p.) drug injections were evaluated. Sodium nitrite (NaNO(2)) and the appropriate solvents served as the positive and negative controls, respectively. Dose-, time-, route-, and compound-related effects were observed. MHb and sulfhemoglobin levels increased, whereas levels of those parameters related to oxygen-carrying capacity of the blood, such as, oxygen saturation and oxyhemoglobin decreased. In general, the effects of PAHP and PAOP were longer lasting than those of PAPP and NaNO(2). Furthermore, PAPP and NaNO(2) were equally effective with either route of administration. Conversely, PAHP and PAOP showed larger effects when administered i.p. versus i.m. The animals treated with N-hydroxy metabolites of the p-aminophenones also showed similar changes in the hematological parameters measured. N-hydroxy PAPP was shown to be the most rapidly acting MHb-forming compound examined in this series. It could achieve therapeutic concentrations of MHb within 2 min and thus may be considered as a treatment for CN intoxication. Although additional work is needed, these data provide information that will be useful for the successful development of improved anti-CN MHb formers.

[Coronary surgery without extracorporeal circulation and Octopus cardiac stabilizer]. / Cirugía coronaria sin circulación extracorpórea y estabilizador cardíaco Octopus.

Between june and july, 1998, surgical myocardial revascularization without cardiopulmonary by-pass through a conventional sternotomy with the use of the "Octopus" heart stabilizer was performed in 6 patients (mean age 63 years, range 49-74 years). All patients received an internal thoracic artery graft. Three of them had also a saphenous vein graft on the distal right coronary artery. An intracoronary shunt was used in four patients and all the anastomoses were accomplished in a completely immobilized area of epicardium. Mean postoperative hemorrhage was 200 ml (50-300 ml) and ICU and total hospital stay were 2 and 6 days respectively. At the first follow-up control, one month post-op, all six patients are free of symptoms without medication. We believe the "Octopus" tissue stabilization system outperforms other heart stabilization devices in our hands and it can be used safely in selected groups of patients.

Psychological casualties resulting from chemical and biological weapons.

This symposium addresses the complications encountered by medical planners when confronted by the use or threat of the use of weapons of mass destruction. The types of chemical warfare agents (CWA), their principal target organs, and physiological effects are discussed. We have reviewed the use of CWA in 20th century warfare and otherwise with emphasis on five cases: (1) use of sulfur mustard during World War I; (2) use by Italy against Ethiopia; (3) use in the Sino-Japanese War; (4) relatively well-studied use in the Iran-Iraq conflict; and (5) the use of sarin in the Tokyo subway terrorist incident. We reviewed the additional physiological and psychological consequences of their use and threat of use. Results from training and simulation are discussed. Finally, we present our conclusions derived from the analysis of these historical situations.

Effects of the organophosphorus compound, O-ethyl-N-dimethyl-phosphoramidocyanidate (tabun), on flavor aversions, locomotor activity, and rotarod performance in rats.

An assessment of the behavioral effects of acutely administered anti-cholinesterase compound O-ethyl-N-dimethyl phosphoramidocyanidate (tabun), to male rats was performed in three studies across five dose levels (100-198 micrograms/kg, sc). Doses of 122 micrograms/kg or greater produced conditioned flavor aversions. Spontaneous locomotor activity was significantly decreased at doses of 122 micrograms/kg or greater when compared to vehicle control levels. When administered doses of 168 micrograms/kg or greater, rats exhibited significant decrements in rotarod performance. In addition, the LD50 of tabun was determined to be 240 micrograms/kg. Signs of cholinergic intoxication were not apparent until 144 micrograms/kg tabun or more were given. Behavioral effects were thus obtained at doses that were between 54 and 71% of the LD50.

Acute behavioral toxicity of the organophosphate sarin in rats.

Three experiments were performed to determine the behavioral effects of acutely administered isopropyl methylphosphonofluoridate, sarin (61-115 micrograms/kg, SC) to male rats. Dosages of 71 micrograms/kg or greater produced conditioned flavor aversions while spontaneous locomotor activity was increased at 61 micrograms/kg and significantly decreased at dosages of 84 to 115 micrograms/kg, when compared to vehicle control levels. Significant decrements in rotorod performance were observed at 98 and 115 micrograms/kg sarin. The LD50 of sarin was determined to be 165 micrograms/kg. Behavioral alterations were thus obtained at dosages that were between 37 and 59 percent of the LD50.

Assessment of motor performance decrement following soman poisoning in mice.

A simple motor performance test, the inverted screen test, was used to assess the incapacitating effects of soman in mice and to evaluate the effectiveness of carbamate pretreatment and/or treatment with atropine plus 2-PAM on soman-induced debilitation. The test requires minimal equipment and personnel training, and can be done rapidly on untrained animals. Mice were placed individually on wire mesh screens which were horizontally mounted on a metal rod. The rod was rotated 180 degrees so that the mice were oriented upside down on the bottom of the screen. The animals were observed for their ability to climb to the top of the screen within one minute. Mice exhibited significant disruption of performance on the screen test 24 hours after soman doses of 0.71 LD50 or more. Treatment with atropine and 2-PAM i.m. 10 sec post-soman (1.1 x LD50) did not improve screen test performance when measured at 24 hours. Pretreatment with pyridostigmine or physostigmine in combination with post-soman therapy with atropine and 2-PAM significantly improved screen test performance as early as 2 hours after soman. The results suggest that the inverted screen test may be useful as a first-line assessment of the efficacy of pretreatment and treatment compounds against nerve agent-induced incapacitation.