Tear Jerker: When D.A. loses her book bag and self-confidence in in "Blows its Top".
Keesha's Heroic BSOD and subsequent ten-minute retirement in "Gets Ants in Its Pants".
The bus almost gets flattened in "Revving Up". The music does not help.
Tough girl Wanda actually cries in "Holiday Special" when her favorite toy from early childhood, which she'd intended to give away to other children who otherwise wouldn't get Christmas presents, gets mulched up and melted down into plastic pellets instead.
Also, in the same episode, the Bus hitting itself with the Recycling Ray, since it has to hit everything that can be recycled, even itself. The fact that it's still smiling just makes the scene all the more unsettling.
The episode wherein Wanda gives up her pet "Bella". For any kid, this can be tough to watch.