Noradrenergic (NA) neurons in the locus coeruleus (LC) play a critical role in behavior by supplying norepinephrine to the brain. Understanding how LC-NA neurons are regulated is essential to understanding the role of the LC-NA system in behavior. Since previous studies have identified a population of inhibitory pre-LC neurons that regulate LC-NA neurons, this study investigates whether there are excitatory pre-LC neurons responsible for the regulation of LC-NA neurons. We infused AAVs into the LC of TH-Cre crossed with vGluT2-FLP mice expressing wheat germ agglutinin (WGA) and eYFP in a Cre- and FLP-dependent manner, respectively. Immunohistochemistry revealed pre-LC neurons with immunoreactivity (IR) to WGA antibody but not to TH antibody. These pre-LC neurons surrounded the LC proper, which contained cell bodies of NA neurons with TH- and WGA-IR. Some pre-LC neurons also showed eYFP signal, indicating that they were glutamatergic; they were mainly located dorsomedial to the LC proper. To characterize their firing properties, we infused AAV carrying floxed GCaMP6f into VGluT2-Cre mice. Brain slices prepared from the mice showed neurons emitting GCaMP6f signal continuously or rhythmically. Cell-attached recording confirmed that they exhibited tonic or rhythmic burst spike patterns, respectively. After cell-attached recording, these pre-LC neurons were recorded in whole-cell configuration and filled with biocytin. The results showed that their axonal buttons contacted the cell bodies or dendrites of NA neurons, indicating functional connectivity between them and NA neurons. In conclusion, this study demonstrates the existence of a population of excitatory pre-LC neurons that also regulate LC-NA neurons.