Navy Captain Jeff Davis said the test, carried out between one aircraft from each side over south central Syria, lasted three minutes and "met its intended objective."
"This test assured that the first time this mode of communication was used would not be during an unplanned encounter," Davis said.
The United States and Russia signed a "memorandum of understanding" on October 20 aimed at de-conflicting their air operations over Syria, where they are waging parallel air campaigns with different objectives.
Over the past two months, the pace of US air strikes has fallen off, compared to the previous six months, hitting a low of 117 air strikes last month.
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The Pentagon denies the drop-off is related to Russia's entry in the conflict, attributing it instead to fluctuations typical of military operations.
US Defense Secretary Ashton Carter last week vowed to intensify coalition air strikes against the IS group, which has taken over large swaths of territory in Iraq and Syria.
"We put on hold all military to military cooperation following the onset of Russian aggression in Crimea, and that remains in effect," the official said, speaking on condition of anonymity.