Isolated segments of guinea‐pig trachea or perfused tracheal tubes were arranged for the recording of trachealis tension changes in Krebs solution containing indomethacin (2.8 μm).
In opened tracheal segments, epithelium removal caused modest (2–3 fold) potentiation of the effects of acetylcholine (ACh) and methacholine (MeCh) but failed to potentiate carbachol (CCh), bethanechol (BeCh), oxotremorine or KCl.
Pretreatment with ecothiopate potentiated effects of ACh and MeCh but not of CCh or BeCh. Removal of epithelium in ecothiopate‐treated tissue potentiated effects of ACh and MeCh but not of CCh or BeCh.
Guinea‐pig ileum challenged with ACh was used as a bioassay system for cholinesterase activity. Scrapings of tracheal epithelium did not hydrolyse ACh.
Histochemical staining revealed no fibres positive for acetylcholinesterase or pseudocholinesterase in the tracheal epithelium. However, the underlying tissues contained acetylcholinesterase‐positive nerve fibres and the trachealis muscle itself stained positively for pseudocholinesterase activity.
Neither tetrodotoxin (3 μm) nor hexamethonium (500 μm) modified the ability of epithelium removal to potentiate ACh.
In perfused tracheal tubes where spasmogens were added to the luminal perfusate, epithelium removal potentiated effects of ACh (31 fold), CCh (10 fold), oxotremorine (2 fold) and KCl.
In perfused tracheal tubes where spasmogens were added to the Krebs solution superfusing the adventitial surface of the tissue, epithelium removal significantly reduced the potency of CCh, oxotremorine and KCl.
It is concluded that the selectivity and magnitude of the potentiation of cholinomimetics caused by epithelium removal depends on the route by which the cholinomimetic agent gains access to the trachealis muscle. The potentiation of acetic acid esters of choline seen in opened tracheal segments does not reflect the loss of epithelial cholinesterase activity and does not depend on the activity of nervous reflex arcs in the tracheal wall. The reduced potency of adventitially‐applied cholinomimetics and KCl seen in epithelium‐denuded tissue strongly suggests that the epithelium can moderate trachealis sensitivity to cholinomimetic agents not only by releasing epithelium‐derived relaxing factor but also by acting as a barrier to drug diffusion.
Citrate inhibited stone growth in this laboratory model. This was true both in defined media and with addition of UMM. This adds to evidence justifying the use of alkaline citrate in calcium oxalate nephrolithiasis.
We have previously shown how individual calcium oxalate stones of about 1 cm can be grown in vitro. While this proved a design concept, it was severely limited as an experimental tool because of the time required to undertake comparative studies. Here we describe a development of this system in which six parallel pairs of stone generators are supplied with feed solutions generating a medium that is supersaturated with calcium oxalate. Twelve stones were grown simultaneously in aseptically prepared artificial urine over a period of 32 days from 100 mg to about 250 mg. Flow rates, pH and [Ca(2+)] were stable and reproducible over the course of the experiment. Sodium azide (0.02%) was included in the growth medium of six stones and caused a modest decrease in growth rate from 5.5 to 3.4 mg/day. The experimental design is such that this was readily detectable both visually and statistically ( p<0.001). This multiple stone growing system ("a stone farm") shows improved consistency and illustrates the statistical power of the technique. Azide has only a minor effect on the growth kinetics and can be used as an antibacterial agent in studies involving urinary macromolecules. The technique is suitable for practical and meaningful investigation of calcium oxalate stone formation in vitro.
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