The US has accused city officials of the Venezuelan capital, Caracas, of complicity in an attack on the car of US Ambassador William Brownfield.
Motorcyclists chased the convoy for at least 10 minutes
The ambassador's convoy was pelted with eggs, onions and tomatoes and chased by motorbikes for some miles by supporters of President Hugo Chavez.
US Under Secretary of State Nicholas Burns said the attack had been condoned by the city government.
However, the mayor's office in Caracas denied any involvement in the incident.
The BBC's Greg Morbasch in Caracas said Mr Brownfield is accustomed to verbal abuse from supporters of the president but this latest incident is the first time he and his team have had objects thrown at them.
Mr Brownfield - who was visiting a low-income neighbourhood in Caracas to donate baseball equipment to underprivileged children - had recently stated he was concerned for his safety.
The US under secretary of state told Venezuelan Ambassador Bernardo Alvarez that if such an incident happens again there would be severe diplomatic consequences, department spokesman Sean McCormack said.
Mr Burns said the attack was a violation of the Vienna Convention and that the action was clearly condoned by the local government, the spokesman said.
Supporters of Hugo Chavez are said to be behind the protests
US Embassy spokesman Brian Penn said the Venezuelan police escorting the convoy did not intervene to stop the incident.
"The motorcyclists were throwing things at us for at least 10 minutes, and the police did nothing... It was serious," he said.
Mr Penn claimed the incident was organised by the mayor's office in Caracas, which has denied any involvement in the incident.
"No official authorised by the mayor's office participated," Luis Martinez, a spokesman for Mayor Juan Barreto, told AP.
Officials said the incident was organised by local residents who wanted Mr Brownfield to leave the area.
Relations between the US and Venezuela have been strained for some time, and Mr Brownfield has faced protests at recent appearances.
The American embassy has also asked the Venezuelan government to improve security for the ambassador, saying it is legally bound to do so.
President Chavez has been at loggerheads with Washington, accusing the Bush administration of orchestrating assassination and coup attempts in order to get at Venezuela's vast oil reserves, our correspondent says.
But US officials say Mr Chavez is causing instability in the region with his fiery anti-Bush rhetoric and autocratic style of leadership.