Being sessile organisms, plants confer various environmental challenges that effect their productivity. Secondary metabolites, including glucosinolate (GSLs), are synthesized in plants to counteract environmental stresses through indirect or direct defensive responses. GSLs are secondary metabolites containing nitrogen and sulfur abundantly found in Brassicaceae, playing pivotal roles in plant stress tolerance. Herein, we delve into emerging insights and unanswered questions regarding the regulation of glucosinolate biosynthesis and their role in stress responses. This review offers comprehensive insights into the diversity and participation of GSLs in sensing osmotic stress and signaling responses during plant development under abiotic stress conditions. Furthermore, we explore the promising research prospects for understanding GSL-mediated stress responses, providing a conceptual foundation for investigating the molecular mechanisms underlying abiotic stress response mediated by GSLs. Future research endeavors should focus on enhancing abiotic stress tolerance in Brassica plants by improving GSL responses through genetic engineering and omics approaches, thereby addressing global food security challenges. It is imperative to note that farmers in developing countries often overlook these methods and resort to agrochemicals to enhance plant growth under unfavorable conditions. Thus, environmentally friendly alternatives to plant-based metabolites, including glucosinolate, must be investigated for improving plant growth under unfavorable conditions using seed treatment or foliar application.