The protesters gathered at a square in front of the Antananarivo town hall on Saturday, calling on President Hery Rajaonarimampianina to quit.
"Get lost, Rajao, murderer, dangerous 'killer'," read one banner.
"We should actually drop the term 'president' from our banners, because for us Rajao is no longer the president of the republic," said opposition lawmaker Lanto Rakotomanga to applause from the crowd, Presstv reported.
The opposition says the government is trying to elbow them out of the race for presidential and legislative elections, due to be held in late November or December, through new electoral laws.
They believe that the new laws could benefit the incumbent and bar some candidates from standing in the presidential election.
Opposition lawmakers have asked the country's constitutional court to examine the new electoral laws and to sack Rajaonarimampianina.
Two people were killed and more than 16 others injured on the first day of the protests last week, which the president denounced as an attempted coup.
Although Madagascar's authorities had announced the protest illegal, some 1,000 opposition activists defied the order and security forces used tear gas to break up the gathering.
The large Indian Ocean island has been beset by decades of political instability. Rajaonarimampianina was elected in 2013 and has not yet announced his decision whether he will stand for re-election.
Marc Ravalomanana, who was president from 2002 to 2009, and Andry Rajoelina, who removed Ravalomanana during a coup, have previously announced that they would contest. The two former heads of state were barred from running in 2013 and their parties have been involved in the demonstrations.
On Wednesday, representatives of both the president's party and that of the opposition met in an attempt to find a solution to the crisis, but to no avail.