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The development of rat intestinal L-Ph specific activity displays a well-known post weaning decline. In contrast, total lactase activity increases to reach maximal levels around weaning, and remains high subsequently. In order to elucidate the molecular basis for these patterns, a rat lactase cDNA was isolated and characterized, and used in the quantification of lactase mRNA during development. This lactase cDNA uniquely hybridized to a 6.8 kb mRNA in the small intestine. To assess the amount of lactase mRNA encoding for lactase enzyme activity in the small intestine, total intestinal RNA was isolated and analyzed by Northern and dot blot hybridization. The pattern of total lactase mRNA during development followed that of total lactase activity, suggesting that over this time span the level of lactase activity is primarily controlled at the transcriptional level. However, the magnitude of increase of total lactase activity during lactation compared to that of total lactase mRNA suggests that additional mechanisms are involved in regulating lactase levels. Analysis of the regional distribution of lactase mRNA along the small intestine at 14 days revealed that mRNA was high in the proximal three regions, but was dramatically lower in the distal regions. Total lactase activity, in contrast, displayed maximum activity in the mid intestine with decreased levels both proximally and distally. Thus, lactase activity in the intestine appears to be regulated during development predominantly by transcriptional mechanisms, while alterations during lactation, and along the proximal to distal gradient, are the result of other control mechanisms.