The India-Pakistan Samjhauta Express bound for Delhi, which was called back from the Wagah border to the Lahore railway station on October 8, left for Attari on Thursday, media reported.
There are 122 Indian and 70 Pakistani passengers on board, officials said.
The train had earlier departed carrying 193 passengers -- both Indian and Pakistani nationals -- but was recalled to Lahore after Indian authorities cautioned Pakistani officials of disruptive farmers' protests in Indian Punjab.
Pakistan Railways (PR) earlier announced that the train would depart on Monday but the departure was delayed a second time as farmers' protests continued on the Indian side of the border.
The train, scheduled to run on Mondays and Thursdays, departed after Indian authorities confirmed resumption of the train service with PR, Pakistan Radio reported.
Indian passengers whose visit visas to Pakistan expired stayed at the PR headquarters and were provided certificates to validate their stay in the country.
Also Read
On Tuesday, 14 stranded Pakistanis who were scheduled to return home by the Samjhauta Express arrived in Lahore from Delhi by the Dosti Bus that runs between the two countries.
Their Indian visas expired and they were to return to Lahore by the Samjhauta train on October 8 and then on October 12 but could not travel because of the cancellation of train service in the wake of farmers' agitation in Indian Punjab.