Vesicular glutamate transporter 2 immunoreactivity in putative vagal mechanosensor terminals of mouse and rat esophagus: indication of a local effector function?

Abstract

Intraganglionic laminar endings (IGLEs) represent the major vagal afferent structures throughout the gastrointestinal tract. Previous ultrastructural investigations have revealed synaptic contacts of IGLEs on myenteric neurons. Thus, in addtion to functioning probably as mechanosensors, IGLEs may also synaptically influence myenteric neurons. In search of clues for potential transmitters in IGLEs, we investigated, by combined neuronal tracing and immunocytochemistry in the esophagus, the correlation between IGLEs and vesicular glutamate transporter 2 (VGLUT2), which is considered a reliable marker for glutamatergic neurons. In rat esophagus, IGLEs were immunostained with calretinin. In the mouse, anterograde wheat germ agglutinin/horseradish peroxidase (WGA-HRP) tracing from nodose ganglion was used in order to label esophageal IGLEs. Confocal laser scanning microscopy demonstrated that VGLUT2 immunoreactivity was highly colocalized with synaptophysin and that both calretinin and tyramide amplified WGA-HRP in rat and mouse esophagus, respectively. No colocalization was found with calcitonin gene-related peptide, a marker for spinal primary afferents. Thus, VGLUT2 is found in vagal afferent endings in the esophagus, suggesting that glutamate is contained in, and probably released from, synaptic vesicles previously described in IGLEs. Functional evidence pending, this finding is in favor of a local effector function of IGLEs onto myenteric neurons.