Louisville is the state's largest city, founded at the Falls of the Ohio in 1778 by Gen. George Rogers Clark. The city was named in honor of Louis XVI of France and that country's aid to the American Colonies during the Revolutionary War. Long a commercial center due to its location on the Ohio River, the city today is a center of health care and medical research (headquartered at the University of Louisville), an international air hub for United Parcel Service, and home of the largest truck-manufacturing plant in the western hemisphere, run by the Ford Motor Company. Historic residential districts include the Victorian Old Louisville, just south of downtown, and the Edwardian Cherokee Triangle to the east. Louisville's Main Street has the largest collection of cast-iron fronted buildings in the United States outside of New York City.

Attractions include historic Churchill Downs race course, home of the Kentucky Derby and the Kentucky Derby Museum; the Belle of Louisville paddle-wheel steamboat, which takes passengers on river excursions; the Louisville Slugger Museum attached to Hillerich and Bradsby Company, where the famous baseball bats are made; a waterfront walk stretching along 16 mi of the Ohio River; a network of parks designed by Frederick Law Olmsted; and historic homes, including Farmington (designed by Thomas Jefferson) and Locust Grove (last home of Gen. George Rogers Clark). Money is currently being raised for a museum dedicated to the life of Louisville's celebrity native, boxer Muhammad Ali. Cave Hill Cemetery, just east of downtown, is a national arboretum. A portion of its 300 acres contains the only federally-funded military cemetery in the country with both Union and Confederate graves, a reflection of Kentucky's position as a border state during the war. Reached from the Indiana shore, but in the river (so still in Kentucky), is the largest exposed Devonian fossil bed in the world, at the Falls of the Ohio.

A rich arts scene is led by the Louisville Orchestra, which gained international prominence in the 1950s for commissioning, performing, and recording works by 20th-century composers. Actors Theatre of Louisville, home of the Humana Festival of New American Plays, which has spawned several Broadway and Pulitzer Prize-winning productions is also at home in Louisville. The city also has resident professional opera, ballet, and children's theater companies. The International Bluegrass Festival is now held in the city each year.