Sussex Police said the incident involved two cars and took place at around 7.25pm.
East Sussex Fire and Rescue Service and the air ambulance were also called to the scene, with the road being closed off to traffic.
Police are now appealing for witnesses following the collision, which took place in Ringmer, near Lewes.
A police spokesperson said: “Emergency services were called to the incident involving a red Peugeot 2008 and a black Audi S3 on the A26 Uckfield Road, outside the Goldcliff Garden Centre at 7.25pm yesterday (May 26).
“Both drivers and both passengers have been taken to hospital following the collision.
“The driver of the Audi, a 29-year-old man from Uckfield, suffered serious injuries.
“The passenger of the Audi, a 23-year-old woman from Uckfield, sustained life-threatening injuries and remains in a critical condition.
“The driver of the Peugeot, a 28-year-old woman from Uckfield, suffered potentially life-changing injuries.
“A 29-year-old man from Uckfield, the passenger of the Peugeot, sustained serious injuries which are not believed to be life-changing or life-threatening at this time.
“Police are investigating the collision and are trying to establish the full circumstances of the incident.
“Officers are appealing for witnesses or anyone with information to come forward. This includes anyone with CCTV in the area, and anyone with dashcam footage from vehicles in the area at the time. Information can be reported to Sussex Police online or emailed to [email protected] quoting Operation Ohio."