The Cherry and Whites had looked destined to seal an historic win in France after tries from Henry Trinder and Charlie Sharples, but the aristocrats of Europe pounced in the dying stages through Clement Poitrenaud.
The build-up to the game had surrounded Mike Tindall's omission from England's Elite squad and £25,000 fine for misdemeanours at the World Cup, but he produced a barnstorming performance and had a hand in both Gloucester tries.
However, it was not enough and the defeat leaves Gloucester third in their pool and with just a losing bonus-point ahead of Saturday's clash with Harlequins at Kingsholm.
Freddie Burns had the first scoring opportunity after just three minutes but missed
a straightforward penalty attempt.
Toulouse quickly found their rhythm and surged into Gloucester territory with some slick offloading.
Yves Donguy appeared to have an open run to the line, but Tindall raced back to make a try-saving tackle.
Gloucester continued to demonstrate their attacking intent and took the lead thanks to the boot of Burns after Toulouse were caught offside at the breakdown on 10 minutes.
Luke McAlister missed a penalty for Toulouse before a deft Burns chip almost paid dividends after 20 minutes, the ball bouncing over Henry Trinder and Tom Voyce while Akapusi Qera was unable to pick it off his toes at full pelt.
But Gloucester were rewarded for their attacking enterprise on 34 minutes.
Tindall received a hospital pass but managed to slip the ball to Trinder before being clattered by Poitrenaud.
Trinder then combined with Jonny May to release Sharples for his third Heineken Cup try. Burns made no mistake with the conversion to put his side 10-0 lead.
But the four-time Heineken Cup winners instantly opened their account through the boot of McAlister before they grabbed a try on 38 minutes.
McAlister threaded a grubber kick through Gloucester's midfield for Timoci Matanavou, leaving Voyce for dead. McAlister missed the conversion.
Gloucester were dealt a major blow 10 minutes into the second half after Alasdair Strokosch was sent to the sin-bin following an indiscretion at the breakdown. McAlister slotted the penalty as Toulouse took an 11-10 lead.
Gloucester were then let off the hook as Donguy lost the ball with a clear run in after Toulouse worked a four-man overlap.
Lionel Beauxis, a replacement for McAlister, missed a drop-goal attempt moments later but stretched his side's lead to 14-10 on 67 minutes.
Gloucester thought they had snatched an historic triumph on French soil after Tindall released Trinder for a vital try.
Burns added the extras to make it 17-14 on 69 minutes but Toulouse hit back almost immediately.
Matanavou danced through Gloucester's defence on the right and offloaded out of contact, the ball flicking off Dave Lewis and into the path of Poitrenaud for the match-winning try which Beauxis expertly converted from the touchline.
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