They have also called on Dumfries and Galloway Council to provide more information to schools and parents about plans to axe more than 50 posts.
A group of parents claim the move could affect all school children, not just those with additional learning needs.
The authority claims changes are needed as part of a drive to save ??32m.
At a meeting in November, the council's education committee agreed to a savings package which would result in the loss of 52 positions from the learning support department.
The petition was set up by parents Roy Holden, of Dumfries, and Demi Powell, of Dalbeattie, who are leading a campaign against the changes.
They both have children who require help from school support staff.
Ms Powell said: "What we are really concerned about is the level of the cuts that are happening. It's a big number of people to lose from the service.
"Parents have struggled to get an appropriate level of support in the current service.
"They are not cutting a service that has a lot of extra capacity. They are cutting a service that's already stretched to the limit."
She said many teachers feel they are not appropriately trained to undertake the work that has been carried out by support staff in the past.
All children will suffer because teachers will have less time to spend with the whole class, she said.
"The teachers have recently implemented a whole new curriculum, and in the secondary schools they have a new qualification system.
"They're being stretched as it is. We want the best for all the children in the region."
A spokesman for the council said that the supported learning department, which has 29 different teams, is "too complex" and could be "significantly improved".
He said the changes will be introduced over three years, it will target those most in need, increase early intervention and it will provide more flexible support tailored to the needs of each child.
A letter is currently being drawn up which will explain the changes to parents and guardians.
He added: "We recognise the need to engage fully with families about the proposals and therefore family engagement forums will shortly be held across the region to ensure that families are kept informed during the development of the new model and to minimise any negative impact for families."
More than 1,000 people have signed an online petition urging councillors to reconsider a decision to cut learning support staff from schools.