Pakistan says its intelligence agents have been accused by the US of alerting al-Qaeda linked militants before the US launches missile attacks against them.
Defence Minister Ahmed Mukhtar said members of Inter-Services Intelligence were accused of "tipping off" militants before strikes in the tribal areas.
Mr Mukhtar said that the Americans "mistrusted" the ISI.
His unusual public admission of the rebuke seems to mark a new low in ties between the US and Pakistan's spies.
'Who's in charge?'
Mr Mukhtar was speaking in Washington, where he is accompanying Prime Minister Yousuf Raza Gilani on his first visit to the US.
Speaking on Pakistan's Geo TV, he said the Americans had alleged that information about targets was being "leaked".
"The burning issue of course is as to who controls the ISI," he said.
The ISI is accused of supporting Taleban fighters in Afghanistan
"In their [the Americans'] view there are some people at some level in the ISI who tip off the Taleban at some level about impending missile attacks when these are shared with the Pakistanis.
"They [the Americans] have expressed displeasure over this."
Mr Mukhtar also openly admitted that President Bush had asked who was really in charge of the ISI during the visit to Washington.
The BBC's security correspondent Rob Watson says that relations between the American Central Intelligence Agency (CIA) and the ISI appear decidedly strained.
Our correspondent says that the stern message apparently being delivered to Islamabad was that Pakistan has to do more to tackle ties between the ISI and Islamic extremists based in the country's tribal areas.
American unhappiness with Pakistan's recently-elected civilian government could also embarrass President Musharraf and the Pakistani army, in addition to raising new concerns about the stability of the nuclear-armed state.
The US no longer gives Pakistan advance notice when it targets militants in tribal areas, correspondents say.
The ISI is the main intelligence wing of the army, which directs its operations, although under the law it reports to the prime minister.
Elements within the ISI have long been accused by politicians and foreign governments alike of supporting the Taleban.
On Saturday, the government said the ISI would be brought under the control of the Interior Ministry.
But the decision was revoked within hours, apparently following intervention from the army.
Alleged US missile strikes have sparked anger in Pakistan
Prime Minister Gilani - whose Pakistan People's Party has a history of run-ins with the ISI - recently called it a "great institution".
He said that he did not believe reports that some members of the ISI were sympathetic to the militants.
On Wednesday, the New York Times reported that a senior CIA official had confronted Pakistani officials with evidence of ISI links to militants, in addition to its involvement in a recent suicide car bomb attack outside the Indian embassy in Kabul that killed 58 people.
Pakistan strenuously denied any links to the attack.
Our correspondent says that while the latest American complaints in relation to the ISI look like something of a low point, it would be wrong to write off the relationship between the ISI and the CIA.
Western security officials often say you cannot deal with violent Islamic extremists without dealing with Pakistan and you cannot deal with Pakistan without the ISI.