Pacific Street Chapel.

The First Presbyterian Church, Newtown (founded in 1652) and the First Presbyterian Church, Jamaica (founded in 1662) were the first organized Presbyterian churches in what is today New York City. Located in the present-day borough of Queens, the original churches faced religious suppression when they were forced to take on Anglican pastors by the ruling Anglican colonial governors. After years of religious persecution, including the imprisonment of the Presbyterian preacher and father of American Presbyterian Francis Makemie, the church eventually was able to expand when the First Presbyterian Church, Manhattan was founded in lower Manhattan in 1716. Soon after, in 1717, another church was organized in Staten Island and in 1822, the First Presbyterian Church of Brooklyn was formed in what is today the neighborhood of Brooklyn Heights.

During the American Revolutionary War, many church members became known for their revolutionary activities against the British. It was also during this period that the church expanded its membership by opening schools and Sunday schools in Manhattan and Brooklyn. Beginning in the 1830s, the issue over slavery eventually led to a split in the Presbyterian Church when the Southern Presbyterian Church was created in 1861. Anti-slavery churches, such as the First Presbyterian Church of Brooklyn faced mob violence during the time leading up to the rupture of the church. Following the Civil War, the church turned its attention to missionary work and social reform. In Brooklyn, the Lafayette Avenue Presbyterian Church sent the first Presbyterian missionary to Korea.

The Presbyterian Church in America faced more challenges during the late 19th and early 20th centuries, including the heresy trial of New York City born theologian and scholar Charles Augustus Briggs. Briggs was one of the early proponents of Higher Criticism, a movement within the church that stressed that the Bible contained errors that should be interpreted as stories and not as facts. This split in doctrine led to what would become the Modernist-Fundamentalist controversy within the American Presbyterian Church, as well as in other Christian denominations in America.

In 1983, the split in the Presbyterian Church that occurred during the Civil War was mended when the church was reunited to form the Presbyterian Church of the United States of America.