They waved flags and carried placards on Wigan Lane on Wednesday morning as their industrial action began.
Nursing staff from more than 55 NHS trusts in England are taking part in industrial action on Wednesday and Thursday following two days of action in December.
The RCN has announced that two further, bigger strikes will be held next month if a resolution is not found.
Pat Cullen, chief executive and general secretary of the RCN, said nurses felt “totally heartbroken” going on strike, but felt they had no choice.
But some services will be affected, with the urgent treatment centre in Wigan only open from 8pm to 1am and the centre in Leigh closed on both days.