Amama Mbabazi was arrested in eastern Jinja town, while Kizza Besigye was detained at his home in the capital, the New Vision newspaper said.
Both are former close allies of President Yoweri Museveni.
He is widely expected to seek re-election next year, but this has not been confirmed.
Last month, Mr Mbabazi said he would challenge Mr Museveni, 70, by seeking the nomination of the ruling National Resistance Movement (NRM) party.
Mr Museveni sacked Mr Mbabazi as prime minister last year, ending a close working relationship of more than 20 years.
Mr Mbabazi was arrested in Jinja as he headed out to canvass support in the eastern town of Mbale, reports the BBC’s Patience Atuhaire from Kampala.
Renegade army General David Sejusa was at the police station in Kampala where Mr Mbabazi was being held to show solidarity with him, she adds.
His daughter has also been arrested, his spokesperson Josephine Mayanja-Nkangi says.
She also says no charges have been brought against him.
On Wednesday, police said Mr Mbabazi had not been cleared to hold meetings.
Dr Besigye, the former physician of Mr Museveni, is seeking the presidential nomination of the opposition Forum for Democratic Change (FDC) party.
Mr Museveni has ruled Uganda since 1986, and his critics say he has become increasingly repressive.
However, his supporters say he has provided stability and has improved the living standards of Ugandans.
By:3news.com/Ghana
Two presidential hopefuls in Uganda arrested
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Two leading presidential hopefuls have been arrested in Uganda for organising meetings without police permission, state media report.