A simple sensitive radioreceptor assay for calcium antagonist drugs

A radioreceptor assay for calcium channel antagonist drugs described here is based on the ability of these drugs to affect ³H-nitrendipine binding to calcium channels. All the known calcium channel antagonists may be assayed in this manner. The assay can detect 10–100 nM (4 – 40 ng/ml) nimodipine, 10–100 nM (3.5 – 35 ng/ml) nifedipine, 3–30 μM (1.2 – 12 μm/ml) prenylamine, 0.1 – 1.0 μM (49 – 490 ng/ml) verapamil and 3–30 μM (1.2 – 12 μg/ml) diltiazem. These values cover the range of concentrations of calcium channel antagonists that are clinically important. As the radioreceptor assay detects active metabolites as well as the parent drugs, it should prove a useful adjunct in cardiovascular therapy. The method is more reproducible, simpler and less expensive than other methods such as high pressure liquid chromatography.     

Extracellular Ca2+ mobilization in potential-dependent contraction of trachealis of maturing swine.

We studied the effect of maturation on potassium-induced parasympathetic activation and Ca2+ entry in tracheal smooth muscle (TSM) from fifteen 2-wk-old (2ws) and sixteen 10-wk-old (10ws) male domestic farm swine. Atropine (10(-7) M) caused inhibition of the maximal contraction elicited by potassium to 50.3 +/- 2.6% maximum of control response (P less than 0.001) in TSM from 2ws but had no significant effect in TSM from 10ws (94.6 +/- 4.2% maximum; P = NS vs. control). Verapamil (10(-7) M) plus 10(-7) M atropine reduced contraction elicited by potassium in both 2ws (23.7 +/- 5.8% maximum; P less than 0.001 vs. control) and 10ws (50.6 +/- 6.3% maximum; P less than 0.001 vs. control, P less than 0.05 vs. 2ws); 10(-6)M verapamil caused greater than 95% blockade of contraction caused by potassium in both 2ws and 10ws. In separate studies, atropine-treated strips were equilibrated with extracellular Ca2+ concentrations ([Ca2+]o) ranging from normal (1X [Ca2+]o) to four times normal (4x [Ca2+]o). Increasing [Ca2+]o increased maximal contractile response in atropine-treated TSM strips from 68.7 +/- 3.8% maximum for 1x [Ca2+]o to 100.8 +/- 4.8% maximum for 4x [Ca2+]o (P less than 0.001) in 2ws. Neither atropine nor [Ca2+]o affected maximal responses of TSM in 10ws (103.5 +/- 3.0% maximum for 1x [Ca2+]o; P = NS vs. control). However, in the presence of atropine and verapamil, 4x [Ca2+]o augmented KCl-elicited contraction of TSM from both 2ws (46.9 +/- 6.3% maximum; P less than 0.01 vs. control) and 10ws (78.6 +/- 2.3% maximum; P less than 0.005 vs. control).(ABSTRACT TRUNCATED AT 250 WORDS)     

Poloxamer 188 decreases susceptibility of artificial lipid membranes to electroporation.

The effect of a nontoxic, nonionic block co-polymeric surface active agent, poloxamer 188, on electroporation of artificial lipid membranes made of azolectin, was investigated. Two different experimental protocols were used in our study: charge pulse and voltage clamp. For the charge pulse protocol, membranes were pulsed with a 10-micronsecond rectangular voltage waveform, after which membrane voltage decay was observed through an external 1-M omega resistance. For the voltage clamp protocol the membranes were pulsed with a waveform that consisted of an initial 10-microsecond rectangular phase, followed by a negative sloped ramp that decayed to zero in the subsequent 500 microseconds. Several parameters characterizing the electroporation process were measured and compared for the control membranes and membranes treated with 1.0 mM poloxamer 188. For both the charge pulse and voltage clamp experiments, the threshold voltage (amplitude of initial rectangular phase) and latency time (time elapsed between the end of rectangular phase and the onset of membrane electroporation) were measured. Membrane conductance (measured 200 microseconds after the initial rectangular phase) and rise time (tr; the time required for the porated membrane to reach a certain conductance value) were also determined for the voltage clamp experiments, and postelectroporation time constant (PE tau; the time constant for transmembrane voltage decay after onset of electroporation) for the charge pulse experiments. The charge pulse experiments were performed on 23 membranes with 10 control and 13 poloxamer-treated membranes, and voltage pulse experiments on 49 membranes with 26 control and 23 poloxamer-treated membranes. For both charge pulse and voltage clamp experiments, poloxamer 188-treated membranes exhibited a statistically higher threshold voltage (p = 0.1 and p = 0.06, respectively), and longer latency time (p = 0.04 and p = 0.05, respectively). Also, poloxamer 188-treated membranes were found to have a relatively lower conductance (p = 0.001), longer time required for the porated membrane to reach a certain conductance value (p = 0.05), and longer postelectroporation time constant (p = 0.005). Furthermore, addition of poloxamer 188 was found to reduce the membrane capacitance by approximately 4-8% in 5 min. These findings suggest that poloxamer 188 adsorbs into the lipid bilayers, thereby decreasing their susceptibility to electroporation.     

In situ treatment of Hamilton Harbour sediment

To enhance the biodegradation of organic contaminants, approximately 18.5 tonnes of oxidant (calcium nitrate) and 5 tonnes of nutrients were injected into sediments of the Dofasco Boatslip, Hamilton Harbour. In the laboratory 78% and 68% of the oil (TPHs) and polynuclear aromatic hydrocarbons (PAHs), respectively biodegraded in 197 days. In the 1992 treatment in the Ddofasco Boatslip, biodegradation of organic contaminants varied from 79% for low molecular weight compounds (BTXs), to 25/15 of the 16 priority pollutant PAHs. At first biodegraduation of large molecular weight PAHs resulted in the production of naphthalene (from 280 g/g to 549 g/g). In the 1993 treatments, 94% of the naphthalene, and 57% of the TPHs biodegraded. The in situ biotreatment of organic contamination takes time but for some sites the significantly lower cost relative to dredging and confinement makes in situ treatment a viable alternative.     

Octopamine receptor subtypes and their modes of action

Octopamine receptor subclasses were first proposed to explain differences in the pharmacological profiles of a range of physiological responses to octopamine obtained in the extensor-tibiae neuromuscular preparation of the locust. Thus, OCTOPAMINE₁ receptors which inhibit an endogenous myogenic rhythm, increase intracellular calcium levels. Also OCTOPAMINE₂ receptors which modulate neuromuscular transmission in this preparation, increase the level of adenylate cyclase activity. The current status of this classification is reviewed by examining the pharmacology of responses to octopamine in a range of preparations. It is concluded that the distinction between OCTOPAMINE₁ and OCTOPAMINE₂ receptor types is still valid, but that OCTOPAMINE₂ receptors exhibit some tissue specific variations. Studies on a clonedDrosophila octopamine/tyramine (phenolamine) receptor are discussed and illustrate many of the difficulties presently encountered in making a definitive classification of octopamine receptors. These include the possibilities that single receptors may activate multiple second messenger systems and that different agonists may differentially couple the same receptor to different second messenger systems.