Variation in lighting environments creates different demands of visual systems for the successful detection and interpretation of visual signals in a given setting. Eye size is a critical property of the visual system as it is has strong effects on visual acuity and visual sensitivity. While many comparative studies have examined eye size across fishes that live in disparate lighting environments (i.e., caves versus surface habitats, mesopelagic versus pelagic depths, turbid versus clear water), fewer have investigated differences in eye size as a function of water clarity at the among population - within species level. Here, we compared relative eye size (eye size residuals on standard length) between wild-caught individuals from swamps and springs and across four drainages in Florida. We also performed a laboratory experiment where we reared animals in clear or tea-stained water, which mimic spring and swamp conditions to determine whether phenotypic plasticity as a function of lighting conditions influences relative eye size. Field caught animals greatly varied in relative eye size among populations, but there was no clear relationship with lighting environment. Fish from southern populations (where swamps are common) had greater relative eye size than those from northern populations (where springs are common). However, in North Florida, bluefin killifish from swamps had smaller eyes than those from springs. This is consistent with the results of our laboratory rearing experiment, which indicated that animals raised in tea-stained water had slightly smaller eyes than those raised in clear water conditions. These two disparate patterns suggest that the determinants of eye size are currently unknown.