Feeding high-grain diets leads to the release and accumulation of short-chain fatty acids in the rumen. The subsequent prolonged decline in ruminal pH can lead to subacute ruminal acidosis (SARA). Accumulation of short-chain fatty acids can cause proliferation of rumen papillae to increase absorption surface, subsequently leading to a thickening of the rumen mucosa. The aim of this study was to evaluate the appropriateness of continuous measurements of the rumen mucosa thickness (RMT) as a diagnostic tool for SARA in dairy cows compared with continuous measurements of ruminal pH. The study used 6 lactating Simmental cows switched from a moderate-grain (MG) diet with 40% concentrate (dry matter basis) for 1 wk to a high-grain (HG) diet with 60% concentrate (dry matter basis) for 4 wk. Reticuloruminal pH was recorded with indwelling sensors throughout the trial. Rumen mucosa thickness was measured by transabdominal ultrasound at 4 d during the MG diet and 23 d during the HG diet. Mean RMT increased from 4.7 ± 0.19 mm in the MG diet to 5.3 ± 0.17 mm in the HG diet, whereas daily mean reticular pH decreased from 6.8 ± 0.01 in the MG diet to 6.5 ± 0.01 in the HG diet. Older cows (>3 lactations) had increased RMT, associated with higher reticular pH throughout the experiment. The higher RMT and pH level in older cows underlines their lesser susceptibility to SARA during high-grain feeding. In conclusion, RMT can successfully be measured using linear ultrasound probes, commonly used by veterinary practitioners as rectal probes. By combining noninvasive RMT measurements with the lactation number of the individual cows in a herd, this study suggests that RMT is a viable option for diagnosing SARA. Further research, using a larger number of cows with different lactations numbers, is needed to establish a cut-off RMT indicating the risk of SARA.