Washington has accused Russia of orchestrating separatist unrest in eastern Ukraine this week and Nato has urged President Vladimir Putin to pull back troops from near the Ukrainian border.
In a statement, the Russian Foreign Ministry said: "The United States and Ukraine have no reason to be worried. Russia has stated many times that it is not carrying out any unusual or unplanned activity on its territory near the border with Ukraine that would be of military significance."
A senior Russian official accused Nato, which restricted its cooperation with Moscow following Russia's annexation of Ukraine's Crimea region last month, of waging a "harsh, uncompromising information war" against Russia and trying to poison its relations with Kiev.
Deputy Defence Minister Anatoly Antonov said: "We see what Nato is doing in Ukraine to prevent them from normalising ties with us.
"Horror stories about the Russian armed forces are being spread."
Nato says Russia has massed tens of thousands of troops near Ukraine's borders and voiced concerns of a repeat of the takeover of Crimea.
The region voted to join Russia in a referendum staged after Russian forces were already in control.
Speaking during a security conference at Russia's top diplomatic university, Mr Antonov said that the turmoil in Ukraine was a result of Western-sponsored efforts to overthrow rulers it disliked and that Moscow considered that any such attempts in the former Soviet Union were aimed against itself.
He indicated Russia was concerned that, since the crisis in Ukraine started, Nato had intensified its contacts with Georgia and Moldova - former Soviet republics that are now eyeing closer integration with the alliance and the West.
The Foreign Ministry also said Moscow has shunned a meeting of the Organization for Security and Cooperation in Europe (OSCE) this week in Vienna, sought by Washington and Kiev over their concerns about Russia's military presence on the border.
The ministry said the US and Ukraine were conducting an anti-Russian campaign at the OSCE and increasing tension over Ukraine, where the authorities are trying to quell unrest by pro-Moscow protesters.
It said: "The everyday activity of Russian troops on its (Russian) territory does not threaten the security of the United States and other OSCE member states. Attempts to accuse Russia of building up its armed forces are groundless."
The turmoil in Ukraine and Russia's annexation of Crimea have led to the biggest East-West standoff since the Cold War.
The European Union said on Tuesday that top diplomats from the European Union, Russia, Ukraine and the United States would meet next week to discuss the crisis.
However, Russian Foreign Minister Sergei Lavrov struck a cool tone and suggested there was no firm agreement.
He said: "Dates have been named (for the meeting) - some time in April.
"But that is what they want. We are awaiting an explanation of what they intend to do at the meeting.
"There are more questions than answers."