Calcium is an important messenger that controls many nuclear functions such as gene expression in mammalian cells but the regulation of nuclear Ca2+ remains unclear. It has long been thought that Ca2+ is translocated from the cytosol by a long distance to the nucleus through the nuclear pore complexes to activate or suppress gene transcription. However, this model is at best an incomplete one. With an aid of confocal and transmission electron microscopy, we demonstrated here that tubules, in a vertical or horizontal orientation, extended deep inside the nucleus of HeLa cells. These nuclear tubules (NTs) are double-membraned invaginations of the nuclear envelope and are usually associated with nucleolus. Also, membrane bound vesicles are found inside and mositol 1,4,5 trisphosphate (IP3) receptors are enriched in some but not all of these tubular structures. Interestingly, shuttling of mitochondria was observed in the NT and cytoplasm of the HeLa cells loaded with dihydro-rhod-2/AM. After stimulation with histamine that increases cytosolic [Ca2+] through IP3 production, a slow rise of dihydro-rhod-2 fluorescence for the measurement of intra-mitochondrial Ca2+ was observed in the area of NT indicating that Ca2+ was sequestered by mitochondria inside the tubular invagination. Our work therefore suggests that the NTs and mitochondrial activities represent a specialized compartment and dynamic process involved in the regulation of Ca2+ inside the cell nucleus. (C) 2003 Elsevier Inc. All rights reserved.