We have imaged sea ice single crystals with X-ray computed tomography, and characterized the thermal evolution of the pore space with percolation theory. Between −18°C and −3°C the porosity ranged from 2 to 12% and we found arrays of near-parallel intracrystalline brine layers whose connectivity and complex morphology varied with temperature. We have computed key porosity-dependent functions of classical percolation theory directly from the thermally driven pore space evolution of an individual sample. This analysis is novel for a natural material and provides the first direct demonstration of a connectivity threshold in the brine microstructure of sea ice. In previous works this critical behavior has been inferred indirectly from bulk property measurements in polycrystalline samples. From a finite-size scaling analysis we find a vertical critical porosity pc,v = 4.6 ± 0.7%. We find lateral anisotropy with pc,pll = 9 ± 2% parallel to the layers and pc,perp = 14 ± 4% perpendicular to them. Lateral connectivity is established at higher brine volumes by the formation of thin necks between the brine layers. We relate these results to measured anisotropy in the bulk dc conductivity and fluid permeability using a dual porosity conceptual model. Our results shed new light on the complex microstructure of sea ice, highlighting single crystal anisotropy and a step toward a realistic transport property model for sea ice based on percolation theory. We present full experimental details of our imaging and segmentation methodology based on a phase relation formulation more widely applicable to ice-solute systems.