Colonel Gaddafi has said that he "cannot surrender" and has called on his supporters to fight on "even if you do not hear my voice" in an audio message sent to a Syrian TV station.
In the message, which was broadcast on the Al-Raai channel Gaddafi said that "the Libyan people are not a herd of sheep, they cannot be defeated".
"Even if you cannot hear my voice, continue the resistance," Gadhafi said.
Speaking on the 42nd anniversary of his rise to power, Gaddafi referred to his opponents as "occupiers" and said that "my voice represents danger to them".
"Stand up to them from city to city, mountain to mountain, valley to valley. It will continue to be a long battle. Libya will turn into a hell. How can the Libyan people surrender?"
Gaddafi also said that his supporters "are not women" and said that they "are going to keep on fighting".
Syrian-based El Raai has frequently broadcast messages from the dictator and his family.
The whereabouts of Gaddafi are currently unknown, though officials from the rebel-led National Transitional Council have said that they believe he is still in Libya.