Denis Pushilin, the self-proclaimed chairman of the pro-Russian Donetsk People's Republic, holds a news conference to announce a planned referendum. Getty Images
DONETSK, Ukraine—The main pro-Russian separatist groups in eastern Ukraine decided on Thursday to go ahead with a referendum on secession set for Sunday, defying an appeal from Russian President Vladimir Putin a day earlier to postpone the vote to facilitate dialogue with the government in Kiev.

Western officials had reacted cautiously to the latest initiative from the Kremlin, which came after Mr. Putin met in Moscow on Wednesday with Swiss President Didier Burkhalter, who is also chairman of the Organization for Security and Cooperation in Europe. The separatist decision threw into further doubt any hopes of easing tensions sparked by his comments, a change of tone that came after weeks of escalation in the region. (Read the latest updates on the crisis in Ukraine.)
In a move likely to add to the unease, Mr. Putin supervised in Moscow what he said in televised comments were previously planned military exercises. Based on official descriptions, the maneuvers were unusually large, stretching across Russia. They included strategic bombers and the launch of ballistic missiles, officials said, in a simulation of a massive retaliation against an enemy after attack.
Pro-Russia rebels wearing gas masks walk inside the city hall in Mariupol, eastern Ukraine, on Wednesday. Reuters

Speaking at the command center, Mr. Putin didn't comment on the separatists' decisions but said Russia "intends to act with those agreements" reached at his meeting with Burkhalter.
Organizers of the independence referendum say if the vote passes they aim to create a new state called Novorossiya, including the Donetsk and neighboring Luhansk regions. Kiev and Western capitals have called the referendum illegal and illegitimate.
"The vote was 100% against," said Denis Pushilin, head of the self-appointed government of the Donetsk People's Republic, after a session of its 78-member governing council. "We are grateful for President Putin's suggestion but we reflect the voice of the people," he told reporters.
The Russian RIA-Novosti news agency said separatists in Luhansk also decided to go ahead with the referendum, according to the press center of the Army of the Southeast, a separatist group
In Kiev, meanwhile, top officials rejected Moscow's demands that the government end its military operation and negotiate with the pro-Moscow activists.
Russia's Defense Ministry accused Kiev of massing 15,000 troops on the border with Russia. There was no immediate response to that claim from Kiev, which has struggled to rebuild its weakened military in the current crisis.
Mr. Burkhalter said the OSCE has drafted a road map for reducing tensions, including a cease-fire, political dialogue and elections. Details of that plan haven't yet been released and officials in Kiev reacted skeptically, complaining that Ukraine had been left out of the talks in Moscow.
Mr. Putin's comments Wednesday were the first apparent sign of the Kremlin trying to pull the conflict back from the brink of a partition of Ukraine. But officials in Washington were immediately critical of the initiative, saying it didn't go far enough.

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In addition to calling on separatists to delay the referendum—a vote Kiev and the West have denounced as illegal—Mr. Putin softened his criticism of the presidential elections Kiev has set for May 25.
He also said Russia had pulled back troops from Ukraine's border that had been deployed there after Kiev began its military operation against the separatists. But NATO chief Anders Fogh Rasmussen said Thursday the alliance had seen no indication of that.
In Kiev, officials said the military operation against separatists would continue even if the referendum is postponed. Acting President Oleksandr Turchynov said the troops are "defending the lives and health of citizens in an antiterrorist operation being conducted against terrorists, saboteurs and other criminals who kill, torture and kidnap our people."
Ukraine's Interior Ministry said two separatist fighters had been killed overnight in the city of Slovyansk, the rebel fighters' center, after they fired on troops at a government checkpoint. There was no immediate confirmation from the separatist side.
Security Chief Andriy Paribiy said that unidentified armed men had attacked border posts in Donetsk and Luhansk overnight along the frontier with Russia. The attacks came from the Russian side, he said. There was no immediate reaction from Moscow to his claim.
Some in Donetsk were dismayed by Mr. Putin's call to postpone the independence vote, which they called a betrayal, but others admitted it is unlikely to succeed given the fighting and lack of broad public support. Separatist leaders flatly rejected calls they lay down their arms, however.
"It's hard to talk about putting down arms and any talks because too many people have died," separatist leader Miroslav Rudenko told Interfax. Dozens have been killed in recent days in fighting between separatists and Ukrainian forces.
Mr. Rudenko also was skeptical about the prospects for the May 25 presidential vote in his Donetsk region. "I don't see any chance to conduct these elections because there are no people among the candidates who could represent the [Donetsk] region and the southeast as a whole," he said.
Pavel Gubarev, the self-appointed leader of the Donetsk People's Republic, who was released from police custody Wednesday in a prisoner swap for kidnapped security agents, said his group would continue taking hostages to exchange in the event Kiev persists in arresting separatist leaders.
What's News: Situation at Ukraine's border with Russian intensifies, foreign governments help to locate missing Nigerian girls and more. Photo: Associated Press
In Luhansk, separatist activists commandeered a World War II-era tank that had been restored for Victory Day parades and drove it to a security-service building occupied by separatists, Interfax reported.
Despite the secessionist moves, a poll released Thursday by Pew Research found that a majority of Ukrainians want the country to remain a single, unified state. In the more pro-Europe west of the country, 93% of those surveyed said they wanted to maintain Ukraine's current borders, compared with 70% of those polled in the east. Overall, 77% of Ukrainians surveyed want Ukraine to remain a unified country. Only 14% of those polled said they think regions should be permitted to secede if they so desire.
The Pew poll also showed the large amount of discontent with the new powers in Kiev. Excluding Crimea, 49% of Ukrainians polled said the new authorities have influenced the country negatively, as opposed to 41% who approved of the new government's actions. The discontent was pronounced in the country's east, where 67% of people polled said the Kiev authorities have negatively influenced the country. 

—Gregory L. White contributed to this article.

Write to James Marson at james.marson@wsj.com