On the bank of a tranquil English river, a young girl named Alice (whom Thomas, Twilight Sparkle and their friends are hanging out with) grows bored of listening to her older sister read aloud from a history book of William I of England. When her sister chastises Alice's daydreaming, Alice tells her cat Dinah and our heroes that she would prefer to live in a nonsensical dreamland called Wonderland. Alice, Dinah, Thomas, Twilight and the others spot a waistcoat-wearing White Rabbit passing by, and Alice and our heroes give chase as he rushes off claiming to be late for an unknown event. Alice, Thomas, Twilight and their friends follow him into a rabbit hole and they all fall into a labyrinth. Alice's dress balloons out and she and the others begin to float. They see the White Rabbit disappear into a tiny door and try to follow, but the door's talking knob advises them to alter their size using a mysterious drink and food (As Alice drinks the "Drink Me" bottle, Thomas, Twilight and the others shrink along with her, but they did not taste the food and stay the way they are right now). Alice, Thomas, Twilight and the others eventually manage to shrink and pass through the door's keyhole and into Wonderland. They meet several strange characters including the Dodo and Tweddledee and Tweddledum who recount the tale of "The Walrus and the Carpenter."
Alice, Thomas, Twilight and the others eventually find the White Rabbit in his house, but before they can ask what he is late for, they are sent to fetch some gloves. Alice eats a cookie and grows into a giant again, getting herself, Thomas, Twilight and the others stuck in the rabbit's house. The White Rabbit, the Dodo, and chimney sweep Bill the Lizard believe Alice to be a monster and plot to burn the house down. Alice escapes by eating a carrot and shrinking down to the size of an insect. Twilight uses a special shrinking spell on herself and everyone else to shrink to the size as Alice's small size. They meet and sing with some talking flowers, but they chase them away upon accusing them of being weeds. Alice, Thomas, Twilight and the others are then instructed by the hookah-smoking Caterpillar to eat a part of his mushroom grow back to their original size. Alice decides to keep the remaining pieces of the mushroom on hand and Thomas, Twilight and the others can keep their remaining pieces on hoove and cab.
Alice and our heroes meet the Cheshire Cat who advises them to visit the Mad Hatter, March Hare and the Dormouse. The three are hosting a mad tea party and celebrate Alice and our heroes' "unbirthdays", a day where it is not their birthday. The White Rabbit appears, but the March Hare and Mad Hatter destroy his pocketwatch and throw him out of the party. Fed up with all the wonderlandians' rudeness and wackiness, Alice, Thomas, Twilight and the others abandon their pursuit of the White Rabbit and decide to go home, but get lost in the Tulgey Wood. Alice breaks down crying while she sings "Very Good Advice" and Thomas, Twilight and the others all felt so very sad at that song, Alice's crying and at the thought that they will never go home now. The Cheshire Cat appears and leads Alice and our heroes into a giant hedge maze ruled by the tyrannical Queen of Hearts and her smaller husband, the King of Hearts. The Queen beheads anyone who enrages her, and invites Alice in a bizarre croquet match using flamingoes and hedgehogs as the equipment.
The Cheshire Cat appears again and pulls a trick on the Queen which she accuses Alice of doing, and Alice is put on trial. Just then, she remembers that she still has the remains of the Caterpillar's mushroom. She eats it and grows to an enormous height which the King claims is forbidden in court. Tired of Wonderland, Alice openly insults the Queen. However, she had hastily eaten both sides of the mushroom and shrinks to her normal size. She, Thomas, Twilight and the others are forced to flee after the Queen orders Alice's execution. Alice and our heroes becomes pursued by most of Wonderland's characters until they finally reunite with the Doorknob, who then tells them they are having a dream, forcing Alice, Thomas, Twilight and the others to wake themselves up. The film ends as Alice, her sister, Thomas, Twilight and their friends head home for tea.
- This movie takes place before Winnie the Pooh and the Care Bear's Adventures in Wonderland.
- When a shrinken Rainbow Dash, Fluttershy, Heckle and Jeckle all tried to get the key up high from the table, they all tried flying, but when that happened, something stopped them from flying and they all stayed on the ground throughout the movie. At the end of the movie, when everyone wakes up from their Wonderland adventure dream, Rainbow Dash, Fluttershy, Heckle and Jeckle reveal that they can still fly.