Increasing evidence indicates a positive effect of probiotics on the nervous system. The objective of this study was to determine if probiotic Lactobacillus rhamnosus GG (LGG) and/or prebiotics polydextrose/galactooligosaccharide (PDX/GOS) can alter the colonic sensitivity in a neonatal rat model of chronic visceral hyperalgesia and to determine whether altered sensitivity is associated with changes in neurotransmitter levels in the brain. Chronic visceral hyperalgesia was induced in rats by intracolonic administration of zymosan for 3 days during postnatal day 14-16 (P14-P16). After weaning (P21), these pups were divided into groups that received either (1) control diet (CD), (2) PDX/GOS, (3) LGG, or (4) PDX/GOS + LGG. These diets were continued until visceral sensitivity was tested at P60. The viscero-motor response (VMR) to graded colorectal distension (CRD) was determined by measuring the electromyographic (EMG) activity from the abdominal external oblique muscles. The levels of neurotransmitters and biogenic amines were quantified in the frontal cortex, subcortex, brain stem, and cerebellum. At P60, the VMR to CRD in the neonatal zymosan-treated rats was significantly higher than neonatal saline-treated rats. In contrast, neonatal zymosan-treated rats that received PDX/GOS or LGG did not exhibit visceral hyperalgesia. The levels of serotonin, noradrenaline, and dopamine were significantly altered in LGG-treated rats compared to other groups. Results document that in rats LGG can attenuate neonatally induced chronic visceral pain measured in adulthood. Prolonged intake of LGG alters some key brain neurotransmitters and biogenic amines that could be involved in pain modulation.