Hearts, Partick Thistle and Stranraer have all been relegated from their respective divisions after the SPFL board "agreed to draw a line under reconstruction talks" after only 16 of the 42 clubs backed plans for a 14-10-10-10 set-up in an informal vote.
It was also confirmed Championship clubs had approved a reduced 27-game campaign, which is scheduled to kick off on October 17.
Clubs later voted to terminate the season and decide placings on a points-per-game basis but Hearts - who were four points adrift - are not prepared to accept their fate.
A club statement read: "This was the final opportunity for kinship to prevail and for Scottish football to stand together in an emergency. It is an unfortunate condemnation of Scottish football that this was not possible.
"We thank those who were open minded, pragmatic and willing to come together to try and reach a fair outcome for all. Sadly, there were too few of us.
"Now that all other avenues are closed, we are left with no choice but to proceed with a legal challenge. The club has tried throughout these last few months to avoid this course of action but we must now do the right thing by our supporters, our employees, our players and our sponsors, all of whom have been unwavering in their commitment and support.
"We can hold our heads up high as we have acted at all times with integrity, common sense and with the best interests of Scottish football at heart.
"We have stated from the beginning that the unjust and unfair treatment of Hearts, Partick Thistle, Stranraer and indeed other clubs cannot be allowed to go unchallenged. While many weeks have been wasted in trying to find a solution, we must now formally challenge this outcome.
"The club can confirm that the necessary steps have been taken to begin this legal challenge."
📝 Club Statement
💬 To our amazing fans we say that we cannot, and will not, sit idly by and watch the decisions made in the past few months further damage Heart of Midlothian FC. Thank you for fighting for us, now allow us to do the same for you.
➡️ https://t.co/yXTVt1Zayh pic.twitter.com/R28a3N4b3I
— Heart of Midlothian FC (@JamTarts)
SPFL chief executive Neil Doncaster had proposed the permanent alteration after three previous attempts at reconstruction failed to garner enough support.
He said: "Whilst a number of clubs were in favour of a new divisional set-up, the support for it was insufficient and we will now move forward with a fixture programme for season 2020-21 based on the current 12-10-10-10 structure.
"Due to the restrictions forced upon us by the coronavirus outbreak, the Championship clubs also voted overwhelmingly to play each other three times next season, rather than four, which enables a later start to the Championship league season.
"Now that we have a confirmed structure for next season, the SPFL's fixturing team will begin work on the Premiership fixture list, which will start on the weekend of 1 August, and the Championship fixture list, which will start on the weekend of 17 October."
Initial talks on reconstruction were shelved when a number of top-flight clubs baulked at plans for a temporary reconstruction, which would have led to mass relegation in two years' time.
Further attempts from Hearts and Rangers to introduce a three-division set-up both failed before the SPFL took the initiative.
The proposal would have resulted in no relegation and divisional runners-up Inverness, Falkirk and Edinburgh City being promoted, while Highland League champions Brora Rangers and Lowland League winners Kelty Hearts would have joined the league after the pyramid play-offs were abandoned.
The SPFL would have pushed for a general meeting in seven days' time if there had been enough support but the numbers fell well short with some clubs not willing to rush permanent change.
The change would have required 11 of the 12 Premiership clubs voting in favour along with at least six Championship clubs and 75 per cent of the league overall.
Partick Thistle, who were two points from safety with a game in hand, are now facing huge uncertainty given no decision has been made on the fate of the League One and League Two seasons.
Additional reporting by PA.