TREATMENT WITH 5-AMINOLEVULINIC ACID AND PHOTOACTIVATING LIGHT CAUSESDESTRUCTION OF PREIMPLANTATION MOUSE EMBRYOS

Objective: To evaluate the direct effect of photodynamic treatment wit h 5-aminolevulinic acid (ALA) on preimplantation mouse embryos in an i n vitro setting.Design: Preimplantation mouse embryos were incubated w ith or without ALA for 5 hours and followed immediately by light expos ure for 0, 5, or 15 minutes. Comparison of the viability and blastocys t formation was made among different treatment groups. Setting: A conv entional laboratory setting with embryo culture facilities. Interventi ons: Female CD1 mice were superovulated with pregnant mare serum gonad otropin and hCG before mating. Four- and eight-cell embryos and compac ted morulae were flushed from the oviducts and incubated with 0, 0.1, 0.5, 1.0, or 5.0 mM ALA for 5 hours. Embryos subsequently were exposed to photoactivating light for 0, 5, or 15 minutes. Main Outcome Measur es: Microscopic assessment of embryo quality at 12 hours and determina tion of the percentage of embryos progressing to the blastocyst stage at 36 or 60 hours. Results: Incubation of embryos with 0.1, 0.5, 1.0, 5.0 mM ALA without light resulted in 87.3% +/- 1.6%, 84.9% +/- 3.4%, 8 1.4% +/- 1.8%, and 82.8% +/- 4.7% of the embryos developing to blastoc ysts, respectively. In the absence of ALA, light exposure for 0, 5, or 15 minutes resulted in 93.8% +/- 2.3%, 92.3% +/- 2.2%, and 85.9% +/- 1.7% blastocyst formation. Combining treatment of ALA at the same conc entrations with light resulted in 33.3% +/- 2.1%, 0.7% +/- 0.9%, 0%, 0 % (5-minute light), 13.3% +/- 1.0%, 0%, 1.6% +/- 1.3%, 0% (15-minute l ight) blastocyst formation, respectively. When gross morphology was us ed to assess embryo viability at 12 hours, similar results were observ ed. Measurement of the fluorescent spectrum of embryos incubated with ALA indicated that protoporphyrin IX had been formed. Conclusion: Phot odynamic ablation of mouse embryos was achieved with ALA under in vitr o conditions. These results indicate that preimplantation mouse embryo s are capable of converting ALA to the photosensitizer, protoporphyrin IX, and are susceptible to subsequent photoablation. A photodynamic e ffect on the embryo may be important to the successful application of this technique to the treatment of human ectopic pregnancy.