The Spend a Penny scheme, run by the Theatres Trust, is handing out a total of Â£125,000 to up to 10 theatres.
There are often complaints that older theatres do not have enough facilities for women, who can spend the entire interval in a queue to use them.
Theatres can apply for up to Â£15,000 to improve toilets for women, including unisex and gender neutral facilities.
British guidelines specifies that for a 500-seat auditorium, eight ladies' toilets should be provided.
"Even though this seems inadequate for audiences whose time for use is strictly limited, there are many theatres which do not even meet this standard," said the Theatres Trust.
"The anxiety caused by long queues, with unappealing conditions often awaiting even the lucky few, meaningfully diminishes the joy of theatre for about half the audience."
The charity will inspect all the theatres that apply to find out which are most in need of the grant.
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A scheme has been launched to improve the standard of women's toilets at theatres across the UK.