The action comes a day after six constables were suspended for "vandalising private vehicles" during Tuesday's protest by farmers in Maval region.
Police inspector Ashok Patil and sub-inspector Ganesh Mane have been suspended, Home Minister R R Patil said in the Legislative Council.
Six police constables -- S S Gengaje, N A Wadge, J H Korde, R B Bhagvat, V B Nagre and M N Dhankar -- attached to Pune rural police were suspended on Thursday.
Farmers were protesting against a pipeline project for supply of water from Pavna dam to Pimpri Chinchwad township when police opened fire leaving three persons dead.
The issue has rocked the Maharashtra Legislature since Tuesday, with the opposition stalling the proceedings and demanding immediate action against the policemen involved in the firing.
In PHOTOS: Bloodshed on the Mumbai-Pune expressway
4 dead in police firing on Mumbai-Pune Expressway
'There was no need for police to open fire'
Maharashtra monsoon session ends amid din over Pune firing
Din in Parliament over Maharashtra firing