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E. coli endotoxin affinity at the nicotinic2 receptor: receptor theory revisited, ilumina nostros occulae
J F Tomera1, S P Kukulka, K Lilford
1Clinical Pharmacology Laboratory, Shriners Burns Institute, Boston, MA.
Abstract:
This paper examines the interaction of endotoxin at the nicotinic2 receptor (ACh-Rpost) with the adenylate cyclase signal transduction system in skeletal muscle. It reports the drug-receptor (DR) dissociation constant (K, 0.05020 +/- .0015 mg/kg) for E. coli endotoxin. It expands on a previous report (1) which presented the relevance of skeletal muscle cAMP as a metabolic indicator and second messenger influenced by endotoxicosis. The use of a murine endotoxic model allowed for the measuring of K for the antagonist. DR affinity measures which relied on the constrained-slope Schild plot measured it. This report assessed dose-response relations of the competitive antagonist dTC (AN) and their modification (AN') by advancing endotoxicosis. The presence of endotoxin at a fixed final concentration (i.e., 7.75 mg/kg) within the body at the end of a two week period caused a rightward shift in the dTC dose-response curve. Endotoxin desensitized cAMP and caused the dTC curve to be shifted rightward. This report differs from typical pharmacological applications where standard agonist curves are obtainable in the presence of an antagonist. This in vivo model did not allow for the measurement of agonist ACh concentration. Therefore, the use of a pair of compounds (i.e., antagonistic dTC and agonistic endotoxin) and their effects (i.e., suppression of active tension and cAMP desensitization, respectively) allowed us to quantitate K. In summary, this report supports the hypothesis linking the nicotinic2 receptor to the adenylate cyclase signal transduction system and illustrates that endotoxicosis perturbed both.
Insights
This study quantifies the drug-receptor dissociation constant for E. coli endotoxin interacting with skeletal muscle nicotinic acetylcholine receptors. Endotoxin desensitizes cyclic AMP and shifts dose-response curves, indicating its impact on this signaling pathway.
Area of Science:
- Pharmacology
- Molecular Biology
- Skeletal Muscle Physiology
Background:
- Skeletal muscle cyclic AMP (cAMP) serves as a critical metabolic indicator and second messenger.
- Endotoxemia, an infection-induced condition, significantly influences skeletal muscle cAMP levels.
- Previous research established the relevance of cAMP in endotoxicosis.
Purpose of the Study:
- To investigate the interaction between endotoxin and the adenylate cyclase signal transduction system in skeletal muscle.
- To determine the drug-receptor dissociation constant (K) for E. coli endotoxin at the nicotinic acetylcholine receptor (AChRpost).
- To assess how endotoxemia modifies the dose-response relationship of a competitive antagonist (dTC).
Main Methods:
- Utilized a murine endotoxic model for in vivo drug-receptor affinity measurements.
- Employed constrained-slope Schild plot analysis to determine the dissociation constant (K).
- Assessed dose-response curves of the antagonist dTC in the presence of varying endotoxin levels.
Main Results:
- Quantified the drug-receptor dissociation constant (K) for E. coli endotoxin as 0.05020 ± 0.0015 mg/kg.
- Observed a rightward shift in the dTC dose-response curve due to endotoxin presence, indicating competitive antagonism.
- Demonstrated that endotoxin desensitizes cAMP and suppresses active tension in skeletal muscle.
Conclusions:
- The findings support the hypothesis linking the nicotinic2 receptor to the adenylate cyclase signal transduction pathway.
- Endotoxemia significantly perturbs both the nicotinic2 receptor and the adenylate cyclase system in skeletal muscle.
- This study provides a quantitative measure (K) for endotoxin's interaction with the nicotinic receptor in a complex in vivo system.