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<p>The extent to which the central highlands of Madagascar were once covered by forests is still a matter of debate: while reconstructing past environments is inherently difficult, the debate is further hampered by the fact that the evidence documenting land cover changes and their effects on carbon and sediment dynamics in Madagascar has hitherto mainly been derived from lake coring studies. Such studies provide an integrated view over relatively large areas but do not provide information on how land-use change affects hillslopes in terms of carbon and sediment dynamics. Such information would not only be complementary to lake inventories but may also help to correctly interpret lake sediment data. Carbon stable isotope ratios (<span class="inline-formula"><i>δ</i>¹³</span>C) are particularly useful tracers to study the past dynamics of soil carbon over time spans ranging from years to millennia and thus to understand the consequences of land-use change over such time spans. We analysed soil profiles down to a depth of 2 m from pristine forests and grasslands in the Lake Alaotra region in central Madagascar. Along grassland hillslopes, soil organic carbon (SOC) content was low, from 0.4 % to 1.7 % in the top layer, and decreased rapidly to ca. 0.2 % below 100 cm depth. The current vegetation predominantly consists of C<span class="inline-formula">₄</span> grasses (<span class="inline-formula"><i>δ</i>¹³</span>C <span class="inline-formula">∼</span> <span class="inline-formula">−13</span> ‰), yet topsoil <span class="inline-formula"><i>δ</i>¹³</span>C-OC ranges between <span class="inline-formula">−23.0</span> ‰ and <span class="inline-formula">−15.8</span> ‰, and most profiles show a decrease in <span class="inline-formula"><i>δ</i>¹³</span>C-OC with depth. This contrasts with our observations in the C<span class="inline-formula">₃</span>-dominated forest profiles, which show a typical profile whereby <span class="inline-formula"><i>δ</i>¹³</span>C values increase slightly with depth. Moreover, the SOC stock of grasslands was <span class="inline-formula">∼</span> 55.6 % lower than along the forested hillslopes for the upper 0–30 cm layer. <span class="inline-formula"><i>δ</i>¹³</span>C values in grassland and forest profiles converge to similar values (within 2.0 <span class="inline-formula">±</span> 1.8 ‰) at depths below <span class="inline-formula">∼</span> 80 cm, suggesting that the grasslands in the Lake Alaotra region have indeed developed on soils formerly covered by a tree vegetation dominated by C<span class="inline-formula">₃</span> plants. We also observed that the percent of modern carbon (pMC) of the bulk OC in the top, middle and lower middle positions of grasslands was less than 85 % near the surface. This could reflect a combination of (i) the long residence time of forest OC in the soil, (ii) the slow replacement rate of grassland-derived OC (iii) and the substantial erosion of the top positions towards the valley position of grasslands. At the valley positions under grassland, the upper 80 cm contains higher amounts of recent grass-derived OC in comparison to the hillslope positions. This is likely to be related to the higher productivity of the grassland valleys (due to higher moisture and nutrient availability), and the deposition of OC that was eroded further upslope may also have contributed. The method we applied, which is based on the large difference in <span class="inline-formula"><i>δ</i>¹³</span>C values between the two major photosynthetic pathways (C<span class="inline-formula">₃</span> and C<span class="inline-formula">₄</span>) in (sub-)tropical terrestrial environments, provides a relatively straightforward approach to quantitatively determine changing vegetation cover, and we advocate for its broader application across Madagascar to better understand the island's vegetation history.</p>