The rallies came a day after trucks carrying NATO supplies rolled into Afghanistan for the first time in more than seven months, ending a painful chapter in US-Pakistan relations that saw the border closed until Washington apologised for an airstrike that killed 24 Pakistani soldiers.
Pakistan faces a domestic backlash, given rampant anti-American sentiment in the country and the government's failure to force the US to stop drone strikes targeting militants and meet other demands made by parliament.
In Karachi, supporters of religious political party Jamaat-e-Islami gathered after Friday
Prayers and marched through the city's main street, chanting against government's decision to reopen supply route for NATO troops fighting in Afghanistan.
Protesters chanted anti-America and anti-government slogans and demanded the supply route be suspended.
“The restoration of NATO supplies is against Muslims. And the people of Pakistan reject this decision and we vow to continue our struggle until the reversal of the decision,” Mohammad Hussain Mehnati, leader of the religious Jamaat-e-Islami (JI) party in Karachi said.
Elsewhere in Islamabad, supporters of the right-wing Islamist Jamaat-ud-Dawa (JuD) party- an organisation widely believed to be a front group for Lashkar-e-Taiba, a militant organisation- staged a similar rally.
Protesters waved JuD flags and chanted “Wage Jihad against America”.
Hafiz Mohammad Shoaib, a leader of the group, has planned a bigger march on July 8 to show their opposition to the government's decision.
“We will force the government to suspend the supply line. We will protest until this supply route is suspend,” he said.
Pakistan closed the routes in retaliation for the US airstrikes in November that killed the two dozen border troops.
The decision to reopen them, after the US apology, marked an easing of strains in the relationship between Washington and Islamabad in recent months.
The Americans have said they did not intentionally target Pakistani forces, but Pakistan disputed that.
Pakistan and the US also differed over how much Islamabad should be paid for trucks to move through its territory.
In the end, they appeared to compromise with the US issuing an apology but paying no extra fees from the 250 US dollars per truck that it was previously paying.