Justin Edinburgh's Newport only returned to the Football League this season after a 25-year absence but they sampled an evening to remember by knocking out 10-man Brighton, who finished fourth in the 2012/13 Championship, 3-1 after extra time.
Tony Mowbray's Championship side, Middlesbrough, threw away the lead to lose 2-1 to underdogs Accrington Stanley while another second-tier side, Mick McCarthy's Ipswich, were also embarrassed in a 2-0 defeat by League One Stevenage.
Dave Jones' Wednesday, meanwhile, were stunned in a 2-1 loss at the hands of their lower league South Yorkshire rivals, Rotherham United, who booked their place in Thursday lunchtime's second-round draw.
Barnsley were another Championship club whose Capital One Cup campaign was in danger of coming to an end at the first hurdle after David Flitcroft's side were taken to penalties by Scunthorpe United on a late night at Oakwell but they ultimately won.
Plymouth Argyle took Birmingham City to extra time but lost 3-2 at St Andrews, and Cheltenham Town staged a comeback to eventually beat Crawley Town 4-3 following an additional half-hour.
There was no repeat of last season's dream route to the Capital One Cup final for Bradford City as they were knocked out in a 2-1 West Yorkshire derby defeat by Huddersfield Town while Sheffield United were eliminated in the first round by Burton Albion for a second successive season.
There were more problems for Wolverhampton Wanderers after back-to-back relegations as they lost 1-0 to Morecambe while Dagenham & Redbridge were denied their first win in League Cup history in a 3-2 defeat by Brentford.
Bongani Khumalo's late goal gave Doncaster Rovers a 1-0 win over Rochdale, and Championship Queens Park Rangers comfortably progressed as Charlie Austin was among the goals in a 2-0 win at Exeter City.
Troubled Coventry City endured another miserable night as captain Carl Baker was sent off after scoring in a 3-2 defeat by Leyton Orient on a night when MK Dons beat Northampton Town 2-1 in normal time.
Charlton Athletic avoided any upset, with summer signing Simon Church scoring twice in a convincing 4-0 win over Oxford United, and Gianfranco Zola's Watford also dodged any shock as Sean Murray inspired a 3-1 victory against Bristol Rovers.
Another Championship side, Nottingham Forest, were too strong for Hartlepool United in a 3-1 victory, and their second-tier foe, Burnley, also showed their superior ability in a 4-0 win at York City.
Forest's East Midlands neighbours, Derby County also reached the second round with Michael Jacobs' goal earning a 1-0 win at Oldham Athletic.
Another Midlands side, Leicester City, were made to work in a late 2-1 win over Wycombe Wanderers on a night when their fellow Championship side, Bolton Wanderers, won 3-1 at Shrewsbury Town.
Walsall left it late to knock out Port Vale 2-1, Southend United eliminated Yeovil Town, and Swindon also went into the second round after beating Torquay United.
Millwall went through after a 2-1 win over London rivals AFC Wimbledon, and there was success for Bournemouth against South Coast foes Portsmouth in a 1-0 win.
In some of the all-League One encounters, Peterborough United enjoyed a late flurry of goals to defeat Colchester United 5-1 while Bristol City picked up a 2-0 success against Gillingham.
However, League One Crewe Alexandra were surprised 3-2 by League Two Bury but fourth division Mansfield were unable to shock Tranmere Rovers, who won 2-0 at Prenton Park.