Structural and Immunohistochemical Studies of CO2-sensitive Brainstem Neurons

Abstract

<p>We identified C02/H+ sensitive and insensitive neurons in rat locus coeruleus (LC) using whole cell patch clamp techniques. The firing rate was recorded in individual neurons in brainstem slices in aCSF equilibrated with 5% C02. Neurons whose firing rate increased >20% in response to 15% C02 were identified as chemosensitive. Neurons were loaded with Lucifer Yellow, imaged using confocal microscopy, and reconstructed in Neurolucida. The soma of chemosensitive neurons were fusiform with an average aspect ratio of 3.14 while the soma of nonchemosensitive neurons were multipolar with an aspect ratio of 1.4. The primary dendrites on chemosensitive neurons were of small diameter (mean 2.3±1.0 11m) while those on nonchemosensitive neurons were larger in diameter (4.4±1.3 11m). The dendritic field surface area of chemosensitive neurons was 3191 11m2, with a volume of 1112 11m3, while in nonchemosensitive neurons the dendritic surface area was 3751 11m2 and volume was 2153 11m3. The dendrites of chemosensitive neurons were longer than those of nonchemosensitive neurons, terminating near the surface of the slice for chemosensitive neurons. These structural findings have implications for the region of the neuron responsible for chemosensitive signaling and will be used in the development of mathematical models. The response of chemosensitive neurons is largely dependent on pH-sensitive K+ channels, which are inhibited by acidification of the extracellular environment. We used mmunohistochemical methods to reveal the regional and cellular distribution of the voltage gated potassium channel subunit, Kv1.4. This subunit was distributed throughout the brainstem but heavy labeling was observed in the LC. Kv1.4 subunits were localized to the soma and proximal dendrites. Not all LC neurons labeled heavily for Kv1.4. Our findings will complement electrophysiological studies aimed at identifying the K+ channel subunits responsible for C02-responsiveness in chemosensitive neurons.</p><p>This presentation occurred at the Wright State University Campus-Wide Celebration of Research, Scholarship and Creative Activities on April 16, 2010</p>