High light and ultraviolet-B radiation (UV-B) are generally considered to have negative impact on photosynthesis and plant growth. The present study evaluates the tolerance potential of three cultivars of Oryza sativa L. (Kanchana, Mattatriveni and Harsha) seedlings towards high light and UV-B stress on the basis of photosynthetic pigment degradation, chlorophyll a fluorescence parameters and rate of lipid peroxidation, expressed by malondialdehyde content. Surprisingly, it was revealed that Kanchana was the most sensitive cultivar towards high light and at the same time it was the most tolerant cultivar towards UV-B stress. This contrasting feature of Kanchana towards high light and UV-B tolerance was further studied by analyzing photosystem (PS) I and II activity, mitochondrial activity, chlorophyll a fluorescence transient, enzymatic and non-enzymatic antioxidant defense system. Due to the occurrence of more PS I and PSII damages, the inhibition of photochemical efficiency and emission of dissipated energy as heat or fluorescence per PSII reaction center was higher upon high light exposure than UV-B treatments in rice seedlings of Kanchana. The mitochondrial activity was also found to be drastically altered upon high light as compared to UV-B treatments. The UV-B induced accumulation of non-enzymatic antioxidants (proline, total phenolics, sugar and ascorbate) and enzymatic antioxidants (ascorbate peroxidase, guaiacol peroxidase, superoxide dismutase and glutathione reductase) in rice seedlings than those subjected to high light exposure afforded more efficient protection against UV-B radiation in rice seedlings. Our results proved that high tolerance of Kanchana towards UV-B than high light treatments, correlated linearly with the protected photosynthetic and mitochondrial machinery which was provided by upregulation of antioxidants particularly by total phenolics, ascorbate and ascorbate peroxidase in rice seedlings. Data presented in this study conclusively proved that rice cultivar Kanchana respond to different environmental signals independently and tolerance mechanisms to individual stress factors was also varied.