In chemical education, many secondary school students experience difficulties in understanding three mutual related meanings of topics, that is, the macroscopic meaning, the microscopic meaning, and the symbolic meaning. As a consequence, student teachers should be prepared carefully to learn how to teach this difficult issue. This article presents a naturalistic case study of the development of eight student teachers' pedagogical content knowledge (PCK) of the multiple meanings of chemistry topics. The student teachers (all M.Sc.) participated in a teacher education program of which the initial phase focused on learning from teaching instead of learning of teaching. They were asked individually to choose and teach a chemistry curriculum topic with a focus on the macro-micro-symbolic issue. Research data were obtained by interviewing the student teachers individually before and after the lessons. The outcomes indicated a development of student teachers' knowledge of teaching difficulties, for instance, too fast and mainly implicit reasoning between macro- and micro-meaning, and a dominant orientation towards the micro-meaning of topics. A development of knowledge of students' difficulties was also indicated, for instance, difficulties in understanding the macro- and micro-meaning of reaction equations. Implications for the follow-up phases of the program are presented.