Biocytin filling of adult gonadotropin-releasing hormone neurons in situ reveals extensive, spiny, dendritic processes

Abstract

Ultrastructural studies suggest that GnRH neurons receive relatively few synaptic inputs. However, these techniques are biased toward the analysis of portions of the neuron containing GnRH peptide. Using acute brain slices prepared from transgenic GnRH-green fluorescent protein mice, individual fluorescing GnRH neurons were identified, patched, and filled with the small-molecular-weight dye biocytin. Cells were subsequently visualized with an avidin-conjugated fluorophore, and their morphological characteristics were analyzed by confocal microscopy. In total, 45 GnRH neurons from seven adult male and eight diestrus female mice were examined. Unexpectedly, we found that GnRH neurons possess remarkably long dendritic processes, in some cases extending over 1000 microm distal to the cell body. The somata and dendrites of all GnRH neurons were decorated with an assortment of spine-like protrusions, including filopodia, in an heterogeneous manner. Overall, GnRH neurons had a mean dendritic spine density of 0.4 spines/microm, with the highest densities found in the first 50 microm of the dendrite. GnRH neurons with dendrites running in a horizontal orientation had significantly (P < 0.05) more spines than dendrites with a vertical orientation. The comparison of male and female GnRH neurons revealed no sexually differentiated characteristics of somal or dendritic spine density. Using a technique in which the full extent of the GnRH neuron can be visualized, we demonstrate here a previously unrecognized GnRH neuron morphology of long dendrites covered in spines. These observations suggest that GnRH neurons are not poorly innervated and that they receive abundant excitatory synaptic inputs.