Fluorescent liquid pyrene derivative-in-water microemulsions.

A fluorescent liquid pyrene derivative with a high fluorescence quantum yield (65%) in the bulk state is reported. With this as the sole oil phase, stable luminescent oil-in-water microemulsions have been prepared. Increasing the loading of liquid pyrene swells the droplets, as detected by small-angle neutron scattering. These larger droplets have a greater proportion of pyrene excimer emission contribution in their photoluminescence spectra, which leads to a red shift in the chromaticity of the emission.

Regulation of adenosine concentration and cytoprotective effects of novel reversible adenosine deaminase inhibitors.

The physiological role of adenosine (Ado) is well known. Although a number of pharmacological attempts have been made to manipulate Ado concentrations in ischemic conditions in different tissues, none have been clinically accepted up to now, mostly due to insufficient elevation of Ado concentrations or unacceptable toxicity. In this study, we evaluated the biochemical and pharmacological actions of several novel erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) analogs to identify new reversible adenosine deaminase (ADA) inhibitors with potential clinical utility. In cell culture experiments, these compounds elevate cellular Ado concentrations under conditions of simulated ischemic stress but very little, if any, under normoxic conditions. Two compounds were selected for study: 9'-chloro-EHNA (CPC-405) and 9'-phthalimido-EHNA (CPC-406), which specifically inhibit ADA in cell-free preparations as well as in intact cells. CPC-405 and CPC-406 do not affect adenosine kinase activity, and they do not affect adenosine transport (influx). CPC-405 and CPC-406 are also more potent than EHNA in elevating adenosine release from human astrocytoma cells and bovine heart microvascular endothelial cells in 2-deoxyglucose-simulated ischemia or under anaerobic conditions. Inhibition of adenosine deaminase by CPC-405 or CPC-406, as well as the 2'-deoxyadenosine toxicity expressed in the presence of these ADA inhibitors, is reversed when the inhibitors are removed by washing the cells. In the isolated rat heart model of ischemia, these novel ADA inhibitors showed enhanced recovery of left ventricular end-diastolic pressure, left ventricular developed pressure, +dP/dtmax and -dP/dtmax. In the rat hippocampal slice model of hypoxia, these compounds also showed neuroprotective effects on CA1 hypoxic injury. In conclusion, these novel ADA inhibitors may represent clinically useful Ado elevating compounds that show cardioprotective, as well as neuroprotective, effects. Also, their potential for immunotoxicity, if any, appears to be transient in nature, representing an important clinical advantage compared with tight-binding ADA inhibitors such as deoxycoformycin.

Imaging of individual normal and regenerating optic fibers in the brain of living adult goldfish.

Retinal arbors in the tectum of living adult goldfish were imaged to determine whether the structural remodelling and refinement that occurs during development continues in adulthood. Individual optic fibers were labelled by making small injections of the lipophilic fluorescent dye DiI into ventral retina and viewing the exposed tectum through a fluorescence microscope equipped with a cooled CCD camera. Arbors were imaged in the living fish every 30-60 minutes for up to 7 hours. Normal adult goldfish showed no evidence of arbor remodelling during this period, though dynamic movements of varicosities present along axon segments were observed. For comparison, regenerating optic fibers were similarly imaged in fish that had undergone optic nerve crush 2-6 weeks previously. In these fish, dynamic structural changes were seen, including branch remodelling, extension and retraction of growth cones, and movement of varicosities.

Corpus callosotomy for intractable epilepsy: seizure outcome and prognostic factors.

We reviewed the outcome of corpus callosal section in 64 adult and pediatric patients to identify factors associated with a good outcome: 48% of patients had a favorable outcome for overall seizure frequency. Improvement was noted in several seizure types and was most likely for drop attacks, particularly in the setting of a unilateral focal cerebral lesion or a true generalized epilepsy of Lennox-Gastaut type. Poor outcomes for drop attacks were more likely if there was associated severe intellectual handicap or bilateral independent spikes on interictal EEG. Complex partial seizures (CPS), most commonly of frontal lobe origin, also responded favorably. The complications of callosal section were usually mild and transient. New focal seizures occurred in only 2 patients and were not as frequent or disabling as preoperative seizures types. A worthwhile improvement in seizure outcome was achieved by completion of the callosotomy in 6 of 10 patients with unsatisfactory results from anterior callosotomy.

Extracellular calcium does not contribute to cryopreservation-induced cytotoxicity.

The possible role of extracellular calcium ([Ca+2]e) in cryopreservation-induced cytotoxicity was tested using Madin-Darby canine kidney (MDCK) cells and a fluorescent multiple endpoint assay. MDCK cells maintained in 2 mM [Ca+2]e and treated with the calcium ionophore, ionomycin, increased their intracellular calcium ([Ca+2]i) as revealed by the calcium indicator dye, Fluo3 and the bottom-reading spectrofluorometer, CytoFluor 2300. The addition of 10 mM [ethylene bis (oxyethylenenitrilo)]-tetraacetic acid (EGTA) to the extracellular medium before treatment with ionomycin blocked this ionomycin-dependent increase in [Ca+2]i. A number of site and activity-specific fluorescent probes were surveyed to determine which indicator dye might best reveal the ionomycin-induced cytotoxic events during this increase in [Ca+2]i. Although most dyes changed their emission profiles in response to calcium, neutral red was found to best reflect the loss of [Ca+2]i homeostasis. The NR50 for a 15-min exposure to ionomycin in the presence of 2 mM [Ca+2]e was approximately 2 microM ionomycin, but ionomycin had little apparent effect on neutral red retention when 10 mM EGTA was added to the extracellular medium. Thus it was clear that an increase in [Ca+2]i could be cytotoxic to MDCK cells and that neutral red could monitor this cytotoxic episode. To test if [Ca+2]e was similarly cytotoxic during cryopreservation, MDCK cells were subjected to cryopreservation in the presence of dimethylsulfoxide (DMSO). In contrast to previous studies, plasma membrane integrity, not lysosomal function, seemed to best correlate with cell survival subsequent to cryopreservation. In addition, decreasing [Ca+2]e had no discernable effect on the retention of plasma membrane indicator dyes, neutral red, or cell survival. It is concluded that a) plasma membrane indicator dyes, not neutral red, might be better indicators of cytotoxicity occurring during cryopreservation; b) DMSO might be toxic to lysosomes during cryopreservation of cultured cells; and c) although [Ca+2]e can contribute to cytotoxicity, the presence of [Ca+2]e might not influence cryopreservation-induced cytotoxicity.

Cellular alterations produced by the experimental increase in intracellular calcium and the nature of protective effects from pretreatment with nimodipine.

The immortalized septal cell line, SN56 B5 G4, generated by the fusion of mouse septal area cells and neuroblastoma cells, was used to determine if nimodipine, an antagonist of voltage sensitive calcium 'L' channels, might act in a neuroprotective fashion when intracellular calcium levels were raised by incubation in ouabain and monensin. Fluorescent indicator dyes and the automated spectrofluorometer, the CytoFluor 2300, were used to analyze specific cellular targets and functions affected by ouabain and monensin and possible protection by prior incubation with nimodipine. Ouabain and monensin were used together to create a time- and dose-dependent toxic episode. Increases in the emission intensity of Fluo3-AM demonstrated that the concentration of intracellular calcium was monotonically increased by increasing levels of ouabain-monensin. The calcein-AM fluorescent probe indicated that there were no changes in plasma membrane permeability during the toxic episode. Lysosomal integrity decreased as indicated by decreases in neutral red retention. The concentration of free radicals increased as shown by the increase in emission intensity of 2',7'-dichlorfluorescein. Nimodipine pretreatment of the cells incubated with ouabain and monensin resulted in apparent protection of lysosomes and a reduction in the level of free radicals. While nimodipine, by itself, produced a small decrease in intracellular calcium, it actually augmented the ouabain-monensin induced increase in intracellular calcium. The data suggest that in immortalized septal cells, (a) nimodipine offers protection to certain of the responses induced by ouabain-monensin, (b) the protection offered by nimodipine may be independent of antagonism of voltage sensitive calcium channels, and (c) that the protective changes can occur at the same time that intracellular calcium is increasing. These latter observations question the hypothesis that the protection against cell death and dysfunction offered by nimodipine is due solely to maintaining calcium homeostasis.

Transnasal stereotactic biopsy of a clivus tumor. Technical note.

Computerized tomography-guided transnasal stereotactic tissue diagnosis of a lytic lesion in the clivus was performed successfully using the Cosman-Roberts-Wells frame, thus avoiding a major craniotomy. The authors recommend stereotaxis as the preferred technique for biopsy in this region.

Behavioral and anatomical consequences of unilateral fornix lesions and the administration of nimodipine.

Male Wistar rats subjected to unilateral fimbria-fornix transection by mechanical knife cut or to sham operations were tested in a water maze and in an open field. Half the animals in each group were treated with either 0.06 mg/kg nimodipine or vehicle, administered i.p. for 7 days, beginning the day of surgery. Animals were sacrificed and brains were processed for acetylcholine esterase (AChE) histochemistry. In the water maze, lesioned rats showed a significant impairment relative to the sham-operated animals. Nimodipine treatment did not improve performance. There were no differences among the groups in the observed frequencies of the open field behaviors of locomotion, hole-poke, rearing and grooming. A significant reduction of AChE-positive cell bodies was found in the medial septal region on the side of the lesion. There were no differences in water maze performance among groups of rats treated with 0.0, 0.5, 1.0, or 5.0 mg/kg nimodipine for 7 days, beginning the day of fimbria-fornix transection, in an attempt to determine any dose-dependent effect of the drug.

Excessive grooming induced by the administration of codeine and morphine.

Previous studies have shown that peripheral injections of the codeinone RX 336-M can induce excessive grooming in rats in an age-dependent fashion. The present experiment demonstrates that codeine, itself, also induces excessive grooming but with apparent equal effectiveness at all ages tested. Because of the structural similarity between codeine and morphine, the effects of intraperitoneally administered morphine were examined as well. Morphine was found to produce a temporal course of excessive grooming quite different from that produced by codeine. Morphine did not affect grooming in the first half (30 min) of the observation period, but accentuated it, briefly, for the next 30-45 min. Intraventricular administration of codeine at doses of 0.3 or 1.0 microgram/3 microliter had no effect on grooming in animals previously shown to demonstrate excessive grooming in response to either dose.

Spontaneous bodily rotations and direction of locomotion at different times after radio frequency lesions at sites in and near the substantia nigra.

Rats were prepared with radio frequency lesions of the dorsolateral or ventromedial regions of the substantia nigra. Other rats were prepared as operated and unoperated controls for each type of lesion. Their behavior was evaluated in an open field at postoperative days 2, 7, 10, and 15. Three types of behavioral changes were observed over time: those noticeable for a brief period, i.e., a few days, after the lesion (rotational behavior), those lasting 7-10 days after the lesion (turning preferences) and those lasting through the end of the experiment that may be permanent (enhanced locomotion). The early effect of the medioventral lesions was pronounced contralateral rotation while the early effect of the dorsolateral lesion was ipsilateral rotation. This effect of the dorsal lateral lesions was reversed on test days 7 and 10. Lesion-induced turning changes associated with forward locomotion were observed on these two test days as well. By 15 days after surgery the only demonstrable effect of either lesion was enhanced locomotion. The results are discussed in terms of various theories of substantia nigra regulation of motor activities.

Toxicological studies on (2"R)-4'-O-tetrahydropyranyladriamycin, a new antitumor antibiotic. Subacute toxicity study in rats.

(2"R)-4'-O-Tetrahydropyranyladriamycin X HCl (THP), a new anthracycline antitumor antibiotic, was administered to Sprague-Dawley rats intraperitoneally for 13 weeks. In rats receiving 0.4 mg/kg/day, piloerection, emaciation, loose feces and thickening of the injection site were evident, and 7 males and 2 females died after week 12. Inferior body weight gain was observed in both sexes starting week 4 approximately 6. The food consumption also decreased. Hematological examination revealed lower counts of total leucocyte and lymphocyte. At termination there were lower spleen, thymus and testes weights, thickening of the walls of the intestine and stomach, gastric ulceration, presence of ascitic fluid, and congestion and thickening at the injection site. Decreases in the lymphocyte populations of the thymus, spleen and lymph nodes were observed microscopically. A decrease in the number of hematopoietic cells in the bone marrow and a degeneration of the germinal epithelium in the testes were also seen, as were gastrointestinal disturbances. These treatment-related effects were mainly confined to rats receiving 0.4 mg/kg/day and to a lesser extent, to rats receiving 0.1 mg/kg/day. The effects on rats receiving 0.025 mg/kg/day were only at the microscopic level. No rats receiving 0.006 mg/kg/day were toxicologically affected.

Toxicological studies on (2"R)-4'-tetrahydropyranyladriamycin, a new antitumor antibiotic. Chronic toxicity study in rats.

(2"R)-4'-O-Tetrahydropyranyladriamycin X HCl (THP), a new anthracycline antitumor antibiotic, was administered to Sprague-Dawley rats (each group 20 rats) intraperitoneally at dosages of 0.001, 0.008, 0.06 and 0.3 mg/kg/day for 53 weeks. Piloerection, loose feces or perianal staining and thin or emaciated build were observed from week 12 in rats receiving 0.06 or 0.3 mg/kg/day. All rats receiving 0.3 mg/kg/day and 3 males and 1 female receiving 0.06 mg/kg/day died or were prematurely sacrificed in the period from weeks 13 to 44. The overall food consumption and body weight gain of both sexes receiving 0.06 or 0.3 mg/kg/day were lower than those of the controls. Hematological examination showed low leucocyte counts, disturbance in neutrophil and lymphocyte number and low erythrocytic characteristics in animals receiving 0.06 or 0.3 mg/kg/day. At necropsy, macroscopic changes were found at the injection site, throughout the gastrointestinal tract and in the male reproductive system in rats receiving 0.06 or 0.3 mg/kg/day. Microscopic examination revealed decrease in the small lymphocyte population in the hematopoietic and lymphoid system in rats receiving 0.008 to 0.3 mg/kg/day. Degeneration of the germinal epithelium of the subcapsular tubules of the testes, gastric ulceration, epithelial adnexal atrophy of the skin and chronic cellulitis and necrosis at the injection sites were also observed. No rats receiving 0.001 mg/kg/day were affected in these respects.