Brazil has declared Venezuela's most senior diplomat in Brazil, Gerardo Delgado, as persona non grata.
The move came days after Venezuela's decision to expel Brazil's own ambassador to Caracas, Ruy Pereira.
Explaining its decision, Venezuela said Brazil had acted illegally in impeaching its former left-wing president, Dilma Rousseff.
On Saturday, Venezuela also expelled Canada's charge d'affaires, accusing him of interfering in internal affairs.
Canada's foreign ministry retaliated on Monday, announcing that ambassador Wilmer Barrientos Fernández, who was already abroad, would not be allowed to return.
Venezuela's charge d'affaires, Ángel Herrera, was also asked to leave.
Relations with Brazil have deteriorated since President Michel Temer took office last year, following Ms Rousseff's dismissal by Congress for fiscal irregularities.
President Nicolás Maduro described her impeachment as "a right-wing coup".
The head of Venezuela's powerful Constituent Assembly, Delcy Rodriguez, said on Saturday that "diplomatic relations with Brazil will not be restored until the government reinstates the constitutional order it has effectively broken".
The Brazilian government said the move showed "once again the authoritarian nature of President Maduro's administration".
Brazil and Canada have both become outspoken critics of Mr Maduro.
They accuse his socialist government of harassing the opposition and violating human rights.
Canada imposed sanctions on senior Venezuelan officials a few months ago.
Canada and Brazil were among many countries critical of Mr Maduro's decision to convene a Constituent Assembly, which effectively replaced the opposition-controlled National Assembly.
The announcement prompted mass street protests, which killed more than 120 people in four months.
The opposition boycotted the poll in July and also held an unofficial referendum in which they said more than seven million Venezuelans had voted against the constituent assembly.
Mr Maduro's six-year term ends in 2019. He is due to run for re-election next year.
Venezuela has one of the world's highest inflation rates and for years has suffered from a shortage of basic goods, including medicines.
The government blames the crisis on an economic blockade led by the United States, as well as a sharp drop in the international price of oil, its main export.