On Sunday, Trump told a conservative conference in Arizona that Panama was charging US ships "ridiculous, highly unfair" fees to use its namesake canal.
After taking charge of building the canal in the early 20th century, the US turned full control over to Panama in the 1970s via a treaty. But this week, Trump said that if the "rip off" did not stop, he would demand the canal be returned to the US – though he did not specify how.
Trump added he did not want the Panama Canal "falling into the wrong hands" and specifically cited China, which has significant interests in the waterway.
"There's a real US national security interest... in controlling its neutrality," Will Freeman, a fellow on Latin American studies at the Council on Foreign Relations, said of Trump's remarks.
"Trump's statement is mostly about that."
China is the second-largest user of the Panama Canal after the US, according to data. It has major economic investments in the country as well.