Background and aims: Although the prevalence of gamma-hydroxybutyrate (GHB) use is relatively low globally, harms related to the drug appear to be increasing. Few existing studies present reliable, representative, population-level data on GHB-related harms. The aim of this study was to investigate trends in acute GHB-related harms within an ambulance database in Australia. Design, setting and participants: Cross-sectional, retrospective analysis of data on all GHB-related ambulance attendances in the state of Victoria, Australia during a 7-year period (January 2012–December 2018). Measurements: Presentations were characterized based on patient demographics, transport to hospital, co-occurring substance use (i.e. GHB only, alcohol, methamphetamine, heroin, benzodiazepine and cannabis) and clinical presentation (e.g. symptoms of anxiety, psychosis, depression). Findings: There were 5866 GHB-related ambulance attendances between 2012 and 2018, with the prevalence rate increasing from 8.8 per 100 000 population in 2012 to a maximum of 21.7 per 100 000 population in 2017. Methamphetamine [odds ratio (OR) = 6.23, P < 0.001] and benzodiazepine-related (OR = 1.43, P < 0.001) co-occurrences; ages between 18–29 (OR = 6.58, P < 0.001) and 30–39 years (OR = 2.02, P < 0.001); and male gender (OR = 1.23, P < 0.001) were significant predictors of GHB-related attendances. Conclusions: There has been a 147% increase in the prevalence of GHB-related ambulance attendances in Victoria, Australia between 2012 and 2019, largely attributable to a growth in the proportions of people using gamma-hydroxybutyrate alone or concurrently with methamphetamine.