Natural reproduction of muskellunge Esox masquinongy has failed in many waters that formerly supported self-sustaining populations. Laboratory studies showed high muskellunge egg mortality associated with high biological oxygen demand (BOD) substrates where hypoxic conditions developed at the substrate-water interface. Measured dissolved oxygen concentration (DO) at the substrate-water interface in muskellunge spawning sites in eight midwestern lakes showed that four lakes had high DO (mean, 6.0-8.4 mg/liter) and little microstratification, and support self-sustaining muskellunge populations. The remaining four lakes showed extreme DO microstratification and hypoxia (mean, 0.4-2.4 mg/liter) at the substrate-water interface. Populations of the latter are almost solely supported by stocking;Qualitative and quantitative observations of spawning areas in the eight lakes showed that suitable spawning area characteristics include low BOD substrates, dense stonewort Chara sp. beds, or reservoirs where water level is lowered annually, resulting in substrate aeration. Reproduction failure is associated with spawning areas characterized by deep organic matter accumulations and dense macrophyte growth. Improvements of spawning habitat to prevent or alleviate DO depletion are among the options available to manage this species;A method to obtain DO measures within 8 mm of the bottom was developed using oxygen-permeable dialysis tubing to obtain a sample, followed by a modification of the micro-Winkler technique. In controlled experiments, results showed that DO within dialysis tubing reach 99% equilibrium (via diffusion) with environmental DO within 3 hours in flowing water and 5 hours in stagnant water;Statistical analysis of ecological information on 117 selected lakes showed that nine variables accounted for 57% of the variability in natural muskellunge reproduction. Conditions identified as most strongly promoting reproduction were limited northern pike abundance, rising springtime water level, high alkalinity, and high shoreline development factor in drainage lake systems. When organized by discriminant function analysis, 58% of the lakes were classified identically to manager-estimated reproductive level, and 91% were classified within (+OR-)1 reproductive level.