Militant Sikh fundamentalists using bullock carts, felled trees and human barricades brought vehicle traffic across the Indian state of Punjab to a virtual standstill today to protest Prime Minister Indira Gandhi's refusal to accept their demands for increased autonomy.

Authorities reported that 20 persons were killed and more than 150 injured in clashes with police, who opened fire in several confrontations during the eight-hour, widely scattered roadblock campaign. The Sikhs suffered the worst casualties since a violent clash last October outside the parliament building here.

The Sikhs carried out the campaign in spite of the arrest yesterday of more than 1,000 activists of the militant Akali party under stiff preventive detention measures. The entire 87-mile stretch of the Grand Trunk Road from the Sikh's spiritual capital of Amritsar to Ludhiana was blocked by Akali protesters, many of whom carried swords and spears and battled police.

Central Reserve Police who were sent to the Punjab from the Indian capital with shoot-at-sight orders to prevent the burning of buses and cars resorted mainly to using tear gas and charges with batons to break up rock-throwing crowds, according to the state-owned news agency, the Press Trust of India.

In Patiala, in the southeastern part of the state, a 20-year-old Sikh was killed when police shot into the grounds of a temple in response to gunfire from inside, according to the news agency. Four buses were set afire near the Dukhiniwarar temple, the news agency said.

A huge bomb blast rocked the center of Amritsar but caused no casualties, apparently because Sikh merchants had shuttered their shops in support of the statewide protest. Authorities said traffic on the Northern Railway was disrupted when barricades were erected near the village of Thathi Kalan in the Ferozepore district.

Akali Party President Harchand Singh Longowal had warned that yesterday's preventive arrests, which he called "the murder of democracy," would inspire even more Sikh protests.

The Akali party, a reform movement founded in the early 1900s to purify the Sikh religion of its Hindu influences, last month rejected attempts by Gandhi to defuse the year-old confrontation, which has presented the prime minister with one of her most serious and potentially explosive domestic crises in the wake of the Hindu-Moslem rioting in Assam state that has left an estimated 3,000 dead.

Gandhi has made concessions to three Sikh religious demands: that Sikh hymns be broadcast over All-India Radio; that the sale of alcohol, tobacco and meat be banned near the golden temple in Amritsar; and that Sikhs be permitted to carry on airliners nine-inch kirpans, the daggers that they wear as religious practice. After a dagger-point hijacking by a Sikh last year, the kirpans were banned, creating a furor.

Akali leaders also are demanding that the government accept a manifesto redrawing the boundaries of three Indian states to create an enlarged Punjab and giving it autonomous powers that no Indian state has except Kashmir.

The Akalis have demanded that the Indian constitution, which the Sikhs refused to sign in 1950, be amended to grant the new Punjab control of all government functions except defense, foreign relations, currency, telecommunications and railways. Similar autonomy was granted to Kashmir to keep that strategically important and contested Moslem state in the Indian union.

Pressured by tribal separatist movements in the turbulent state of Assam and elsewhere in far northeastern India, Gandhi is viewed as being unlikely to yield to most of the Sikhs' political demands out of fear of encouraging further regionalism in this culturally, ethnically and linguistically diverse country.

But she has come under criticism from some opposition leaders for not having made religious concessions before tensions began to increase in the Punjab and before the Akali movement gained momentum.