Hi, i want to get U46619 concentration-response curve in basilary artery of sprague-dawley rats. I made a stock dilution 2e-3 mol/L in ethanol with 1mg U46619 (Cayman Chem. 1mg in 100 mikroliter methyl acetate).Then in every experiments, i make a series of dilution 2e-3, 2e-4, 2e-5, 2e-6, 2e-7. So i get the cumulative responses for concentration 1e-10 to 3e-6 mol/L in bath. I have a bath with 5 mL in wire myograph. There are some problems:
1) U46619 constricts the artery suddenly, nearly to maximum.
2) U46619 responses have a great variability. So concentration response curve has a great SEM.
3) The amount of ethanol that is given to bath is important? If i reduce the amount of ethanol, does the response change?
Do you have an idea to get more stable U46619 contrations?
Maybe you can find a clue in these articles
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2707971/
http://ajpheart.physiology.org/content/ajpheart/284/2/H719.full.pdf
I haven´t worked with basilar arteries, but mesenteric artery seems to behave quite similar to what you explained, giving sudden contractions, that usually are not stable over time. There is no much to do about it, although I replaced Krebs with DMEM solution and I was somehow able to improve the stability of the responses (no FBS in the medium, as it would give foam).
Regarding the amount of ethanol in your bath is surely important. Is it necessary to make every dilution coming from the original stock in ethanol? If you were to use water in the lower dilutions you would be adding less solvent into your bath. As a matter of fact for DMSO people avoid to go over 0.1% concentration. Being the myograph bath 5 ml in volume we would be talking of 50microL. With the range of concentrations you are applying I don´t think you would be adding more than that volume. Regarding ethanol, I am not sure what is the volume you can tolerate into your bath, but you could try DMSO instead.
I hope these ideas can help you.
Good luck.
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