The aim of the present study was to determine whether pharmaceutical preparations with pregabalin (PGB) as an active ingredient, which are widely prescribed by clinicians, exert toxic effects on human primary nucleus pulposus (NP) and annulus fibrosis (AF). Primary human cell cultures were obtained from intact (n=6) and degenerated (n=6) tissues resected from the two groups of patients. Different doses of PGB were applied to these cultures and cells were subjected to molecular analyses at 0, 24 and 48 h. Cell vitality, toxicity and proliferation were assessed using a spectrophotometer. The expression of chondroadherin (CHAD), a (member of the NP-specific protein family), hypoxia-inducible factor-1 alpha (HIF-1 alpha) and type II collagen (COL2A1) was measured using reverse transcription-quantitative polymerase chain reaction. The results revealed that cell intensity increased in a time-dependent manner and cell vitality continued in the cultures without pharmaceuticals. Cell proliferation was suppressed in the PGB-treated cultures independent from the dose and duration of application. PGB was demonstrated to suppress the expression of CHAD and HIF-1 alpha. In contrast, COL2A1 gene expression was not revealed in any experimental group. The present study utilized an in vitro model and the PGB active ingredient used herein may not be representative of clinical applications; however, the results demonstrated that PGB has a toxic effect on NP/AF cell cultures containing primary human intervertebral disc tissue. In summary, the use of pharmacological agents containing PGB may suppress the proliferation and differentiation of NP/AF cells and/or tissues, which should be considered when deciding on an appropriate treatment regime.