[en] A rat hepatocyte cell line was shown previously to contain a nuclear pool of free heparan SO₄ (HS) chains which turns over rapidly. These cells were labeled with ³⁵SO₄^2- and the steady state levels of [³⁵SO₄]HS in the nucleus were measured under culture conditions chosen to give different cell doubling times (DT's). In each log phase culture growing at a constant rate, the level of nuclear HS did not change as the cell density increased. Rapidly dividing cells had levels of nuclear HS within the range of 30-42 pmol ³⁵SO₄ in the nuclear HS/10⁶ cells. However, in cultures with decreased DT's, there was a corresponding decrease in the level of nuclear HS below 30 pmol/10⁶ cells that was proportional to the decrease in the DT. In contrast, when the cell DT increased in cultures approaching confluence, there was a corresponding increase in the level of nuclear HS above 42 pmols/10⁶ cells. The level of nuclear HS in cells which showed constant inhibition was maximal at 65-90 pmol ³⁵SO₄ in HS/10⁶ cells. Serum, insulin, hydrocortisone, and retinoic acid increased the amount of nuclear HS in log phase cells, whereas phorbol-12-myristate-13-acetate, diBucAMP, and decreased levels of inositol or choline lowered the amount. The correlation between DT's and nuclear HS levels suggest a role for nuclear HS in the regulation of cell division