Worm Breeder's Gazette 15(3): 9 (June 1, 1998)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
1012 Fairchild Center, Columbia University, New York, NY 10027
As noted by Dent & Avery (WBG 15(2):16), drugs used to paralyze worms either quench GFP fluorescence (Na azide) or create unwanted distortions in morphology (levamisol, 1-phenoxy-2-propanol, ethanol). At the suggestion of Micah Siegal (California Institute of Technology), we tested whether 2,3-butanedione monoxime (BDM, Aldrich Cat #11, 213-5), a drug that interferes with stimulus-contraction coupling in vertebrate muscle , could paralyze intact worms. We find that BDM is effective for paralyzing worms and inhibiting pharyngeal muscle contractions. Both GFP fluoresence and worm morphology (apart from the occasional vacuole) are preserved during BDM treatment. Upon exposure to a buffered solution of BDM (0. 3M BDM, 10 mM NaHEPES, pH 7.2), worms begin to twitch and eventually become paralyzed. Worms readily recover from brief (< 20 min) exposures and eggs from treated adults appear to hatch and develop normally. The effect of BDM is concentration-dependent: 83 +/- 8% (mean +/- s.d., n = 360) and 39 +/- 2% (n = 345) of worms are paralyzed after 10 min in a buffered solution of 0.3 and 0.1 M BDM, respectively. While high concentrations of BDM are needed to paralyze intact worms within a reasonable amount of time, the effect cannot be purely osmotic since soaking worms in physiological saline of comparable osmolarity (323 mOsm) does not result in paralysis. Since BDM is effective at millimolar concentrations in vertebrate muscle, we believe that high concentrations are needed to paralyze intact worms because of the barrier presented by the cuticle. Our current procedure for using BDM is: (1) wet a small filter disk (0.25" diameter, punched out of Whatman No. 2 filter paper with a hole punch) with 6 microliters of 0.3 M buffered BDM, (2) to transfer worms in ~4 microliters of M9 to the surface of the filter disk, (3) incubate for 10 min at room temperature, (4) invert the filter disk on the surface a 2% agarose pad (in water), remove filter and gently apply a coverslip. Worms can also be paralyzed by transferring them to the surface of a BDM-agarose pad. A second layer of agarose can be used in place of the coverslip and may reduce the possibility of crushing worms.