Functional properties indicate high-density host engraftment by both human glial progenitors and astrocytes(A) Large and symmetric EGFP+ cell (green) in an acute cortical slice prepared from a mouse engrafted with human glial progenitors 4 months earlier. Inset: lower magnification of the same field. The EGFP+ cell was loaded filled with Rhod2 (red) by a path pipette. Rhod2 diffused into several neighboring EGFP- cells (white arrows, top panel). Cell identity was verified by immunolabeling against GFAP (red, below panel). Neighboring cells were GFAP+ and their shape characteristic of mouse astrocytes indicating that the human EGFP+/GFAP+ astrocytes was couple by functional gap junctions to host GFAP+ astrocytes. (B) I-V curves from host mouse astrocytes n=17; smaller, less complex EGFP+ cells, presumably glial progenitor cells, n=14; and large and symmetric EGFP+ cells, presumably astrocytes, n=37. (C, D) Comparison of the input resistance and gap junction coupled cells detected as number of neighboring cells labeled with Alexa 594. Mouse and large EGFP+ cells (presumed human astrocytes) have a low input resistance, and are extensively coupled by gap junctions. In contrast, small EGFP+ cells - presumed human GPCs - exhibited high input resistance and were not gap junction coupled. n=14-37, *; p < 0.05, Steel-Dwass test. (E) Photolysis of caged Ca2+ in an EGFP+ astrocytic process. white “X” shows initiated point; white arrowhead shows Ca2+ propagation. (F) Top panel, line scan position across the length of a mouse astrocyte filled with NP-EGTA and rhod2. Below, line scan image of an intra-astrocytic Ca2+ wave initiated by photolysis of the cell body. White dashed line indicates the velocity of the intracellular Ca2+ wave. (G) Line scan image of a human astrocyte in a chimeric mouse (H) Comparison of velocities of intracellular Ca2+ waves in host murine and engrafted human EGFP+ astrocytes, and in human astrocytes in freshly resected surgical tissue. n=8-35, *; p < 0.05, Steel-Dwass test.Scale: 30 μm (A); 100 μm (A) insert; 20 μm (B); 10 μm (E). Data graphed as means ± SEM.
