This study investigates the interlayer shear performance of 3D-printed concrete (3DPC) using direct shear tests. Three nominal layer heights, 5 mm, 10 mm, and 15 mm, were considered, and specimens were loaded parallel to the printing path (x direction) and perpendicular to the printing path (y direction). The results show that the interlayer nominal shear strength decreased with increasing layer height. When the layer height increased from 5 mm to 10 mm and then to 15 mm, the nominal shear strength decreased from 9.18 MPa to 7.01 MPa and 4.88 MPa in the x direction, and from 7.87 MPa to 5.29 MPa and 2.68 MPa in the y direction. At the same layer height, the x-direction specimens exhibited higher nominal shear strength than the corresponding y-direction specimens, with increases of approximately 17%, 33%, and 82% for the 5 mm, 10 mm, and 15 mm series, respectively. DIC analysis indicated that tensile–shear damage was the main local failure characteristic. The loading-direction effect was related to different shear-transfer paths: the L-x specimens mainly followed a “continuous filaments-mortar matrix-interlayer bonding” path, whereas the L-y specimens were more controlled by weak interlayer-edge regions and local stress concentration. The effective shear-area analysis showed that the effective bonded area decreased with increasing layer height. After area correction, the corrected shear strengths of the x-direction specimens were 9.18 MPa, 8.76 MPa, and 8.13 MPa for L-5-x, L-10-x, and L-15-x, respectively, while those of the y-direction specimens were 7.87 MPa, 6.61 MPa, and 4.47 MPa, respectively. This indicates that a larger layer height not only reduced the effective bonded area but also weakened filament compaction and bonding quality. The findings provide a mechanism-oriented basis for understanding the anisotropic interlayer shear behavior of 3DPC.