Basically, another way of saying "Shut up and take it like a man" or "Quit your whining." The term is derived from the popular image of cowboys being tough, unflinching, uncomplaining, and hard-working.
"Cowboy up, man. What would John Wayne say if he heard you complaining over a little hard work?"
by You Can't Kill the Metal October 06, 2006
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Another incredibly vacuous term that no longer has the same meaning it initially did, thanks to cultural theft and abuse by people who aren't really cowboys at all, but bored, middle-class white people. It used to mean to toughen up or approach a problem with a hard attitude, but now is used to refer to anything requiring effort, or anything in which people are competeing against each other.

People sport the logo on their clothing or their car because they think it gives them credibility as a cowboy or a redneck, since it is now cool to be associated with these groups once again. But when you get down to brass tacks, most fans of the cowboy up marketing scheme are those who have no connection with the cowboy way of life at all. See poser, git er done and white trash.
Jenny thought that putting her "cowboy up" sticker on her SUV would make others think she was a country girl. In reality, most people just laughed at her.
by Eric in NE March 01, 2006
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A term used by small city fire departments when a Big City Fire Fighter takes initiative to accomplish a task during an emergency without being told to do so.
Did you see the Big City Fire Engineer "Cowboy up" when he took lights and blowers to the front door of that structure fire? I.C. never requested that.
by Johnny Wayne October 05, 2013
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Used in the same sense as the expression "get yourself together." Often heard with an expletive.
"Cowboy the f*** up!"
by Sage November 16, 2004
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What your fat hillbilly neighbor says when he's ready to get going.
You: We ready to go mow some lawns now?
RJ (typical hillbilly name): YEAH BOY, LETS GO BABY, Cowboy Up!!!!!!
You: Ok then...
by Lord_Ahrim1536 June 25, 2015
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Phrase used to describe to action of taking a deep breath, forgetting about the pain and fear, digging down deep, and carrying on to the end.
Dad: Son, are you injured or are you hurt?

Son: What's the difference?

Dad: If you are injured, you sit your ass on the sidelines and watch your team play the rest of the game. If you are hurt, you cowboy the fuck up, swallow your pain, put your helmet back on, and get back into the fight.

Son: I'm just hurt a little, dad.

Dad: That's my boy.
by dammit janet October 21, 2004
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