Gibraltar says it will not be used as a bargaining chip in negotiations over Brexit.
Its chief minister Fabian Picardo says the message that “we want to stay British” needs to be clear in every capital of the European Union.
In draft Brexit negotiating guidelines, the EU said any decisions affecting Gibraltar would be run past Spain.
Theresa May has said the UK is “committed” to the territory and its sovereignty is not on the table.
Gibraltar has accused Spain of manipulating the European Council for its own political interests.
Mr Picardo said: “Let us be very clear and let the message be clear in Madrid, in Brussels and in every other capital of the European Union.
“Gibraltar is not a bargaining chip in these negotiations. Gibraltar belongs to the Gibraltarians and we want to stay British.”
The EU’s guidelines followed a letter from Mrs May formally triggering Brexit talks, which did not mention Gibraltar directly.
After Mrs May spoke to Mr Picardo on Sunday, Downing Street said: “The prime minister said we will never enter into arrangements under which the people of Gibraltar would pass under the sovereignty of another state against their freely and democratically expressed wishes, nor will we ever enter into a process of sovereignty negotiations with which Gibraltar is not content.
“The prime minister said we remain absolutely dedicated to working with Gibraltar for the best possible outcome on Brexit and will continue to involve them fully in the process.”
Mr Picardo said he was “energetically and enthusiastically” supporting Mrs May in her Brexit negotiations.
Gibraltar: key facts
- Gibraltarians are British citizens but they run their own affairs under a chief minister
- The territory is self-governing in all matters – including taxation – except foreign policy and defence, which are dealt with by the UK government
- Despite its small size, Gibraltar is strategically important, standing only 12 miles from the north coast of Africa. It has a UK military base, including a port and airstrip
Defence Secretary Sir Michael Fallon told the BBC on Sunday that the UK would protect Gibraltar “all the way” because its residents had “made it very clear they do not want to live under Spanish rule”.
Former Conservative leader Lord Howard has suggested the prime minister would defend Gibraltar in the same way as Margaret Thatcher defended the Falklands.
He told the BBC Theresa May would show the same “resolve” as Mrs Thatcher did over the 1982 conflict.
Lord Howard said that 35 years ago, “another woman prime minister sent a taskforce halfway across the world to protect another small group of British people against another Spanish-speaking country.
“And I’m absolutely clear that our current woman prime minister will show the same resolve in relation to Gibraltar as her predecessor did.”
After Argentina invaded the Falklands in 1982, Margaret Thatcher sent a task force to reclaim the islands, in the South Atlantic.
An estimated 655 Argentine and 255 British servicemen lost their lives in the fighting that followed.
Labour’s shadow foreign secretary Emily Thornberry said Lord Howard’s comments were “inflammatory” and would “not help Britain get what it needs from these difficult Brexit negotiations”.
Lib Dem leader Tim Farron added: “In only a few days the Conservative right are turning long-term allies into potential enemies.
“I hope this isn’t a sign of the government’s approach to the long negotiations to come.
“Brexiteers have gone from cheering to sabre-rattling for war in four days, it is absolutely ludicrous.”
Spain has long contested Britain’s 300 year-rule of Gibraltar.
Gibraltarians, who number about 30,000, rejected by 99% to 1% the idea of the UK sharing sovereignty with Spain, in a vote in 2002.