In vivo neural activity measured with FM 1-43 in rat mesenteric microcirculation

To date, local neural control of microcirculation has been investigated in an indirect manner due to inability to visualize or directly record activity of small terminal nerve fibers in vivo. Thus, the present study was designed to measure neural activity associated with intact mesenteric microcirculation in vivo. First, in order to determine the location of sympathetic microcirculatory innervation, we employed dopamine-β-hydroxylase (DBH) immunofluorescence in paraformaldahyde fixed rat mesenteries. Most DBH-positive fibers were noted to course along with microvessels down to diameters as small as 8 μm. Nevertheless, not all microvessels were associated with DBH-positive fibers, and visa versa. To visualize neural activity associated with mesenteric micorcirculation in vivo, the fluorescent dye FM 1-43 was employed. Rats were anesthetized with Inactin and their mesentery was exteriorized for viewing of the microcirculation. The mesentery was supervised with 10 μM FM 1-43 for 2 minutes followed by a 4 minute wash with Ringers and 4 minutes monitoring of FM 1-43 fluorescence decay. This procedure was repeated following 2 minutes exposure to 1% lidocaine. The pattern of FM 1-43 fluorescence was similar to the pattern of DBH staining, indicating uptake of the dye by sympathetic neurons. FM 1-43 fluorescence decay rate is proportional to the rate of neural exocytotic activity. Prior to lidocaine exposure, FM 1-43 fluorescence at single neural boutons decayed rapidly. This decay was significantly attenuated by lidocaine. Thus, FM 1-43 fluorescence may be used to monitor neuronal exocytotic activity in an in vivo microcirculatory preparation.