The conservative Popular Party government of Valencia opened the investigation after children at the Taquígrafo Martí primary school in the town of Xàtiva booed regional president Alberto Fabra on December 4th.
The hostile reception took place in an atmosphere of social unrest and anger following a decision by his government to shut down the regional television and radio network RTVV — the only Catalan language media outlet in the region.
"We love Channel 9" and "Bring back Channel 9", the children of the predominantly Catalan-speaking school shouted.
In the wake of this protest at the school, the regional government launched an investigation to see whether children had been forced to demonstrate by their teachers.
It was "shameful" to "manipulate young children," said Valencia's minister for governance and justice Serafín Castellano.
But teachers at the school told Spanish news agency Europa Press the demonstration was spontaneous. They said it was "coincidence" that Fabra had been near the school during play time.
Regional teachers union the STEPV called the investigation "authoritarian".
On Monday, however, regional education minister María José Català said the investigation was going ahead. The plan was not to "punish" the school, she said, but to find out what had taken place.
"We are going to analyze the situation and, of course, we won't consent to, or tolerate, any use of children for purposes which aren't part of school activity," said Català.
The STEPV said this was about "instilling" fear among teachers and said schools should demonstrate where they saw fit.
In another incident in September, Spanish politicians expressed their outrage over a television programme during which they said 'brainwashed' children from Catalonia had expressed pro-independence views.