Rationale: Phenylethanolamine N-methyl transferase (PNMT) is the enzyme that catalyses the conversion of noradrenaline into adrenaline in endocrine cells. Our group previously reported a new type of cardiomyocytes that historically express PNMT during development stage. We call these cells as Pnmt-derived cardiomyocytes (PdCMs)1. PdCMs have unique left heart localisation in adult heart1. However, so far little is known about their molecular signature and their potential additional function in physiological or pathophysiological conditions comparing to conventional cardiomyocytes due to their neuronal origin. . Methods: single cardiac cells were isolated from either C57BL neonatal mouse hearts (aged 3 days, n=20) or adult (12 weeks old) YFP-ChR2/Cre-Pnmt mouse hearts for Single-cell RNA sequencing (scRNAseq). For neonatal cells, 10X Genomics scRNAseq was used. Single cell suspensions were converted to barcoded scRNA-seq libraries with the Chromium Single Cell 3′ kit with 10X Genomics platform, aiming for an estimated 10,000 cells per library. The libraries were sequenced using HiSeq400. Expression matrices were generated using CellRanger and analysed by Seurat package. Dimensionality reduction using principle component analysis was applied to identify major cell types and their subtypes. For adult myocytes, SORT- RNAseq was used. Results: In neonatal heart samples, a total of 10550 transcripts were sequenced at single cell resolution. Major cell types were identified by leveraging single-cell transcriptomics analysis. We found Pnmt expressing cells are exclusively cardiomyocytes in neonatal heart. Rspo3 and Bmp2 are the most differentially expressed genes in Pnmt expressing cardiomyocytes compared to conventional cardiomyocytes. The clustering of Pnmt expressing cells are mainly presented in cardiac conduction system (CCS), left atrium and left ventricle. In adult mice, 192 YFP-Pnmt negative and 192 YFP-Pnmt positive cells were sequenced. Expression of Pnmt gene was significantly declined and even disappeared in most PdCMs. However, in both neonatal and adult Pnmt+ cells, a high expression of exocytosis-related genes including CPLX2, SLC12A2, SLC6A4, SLC22A1, RAB31, RAB35, RAP2B, STX7, VAMP3, VAMP4 have been identified. Conclusions: Single-cell RNA sequencing revealed molecular signature of Pnmt-derived myocytes. Their high expression of exocytosis-related gene expression and special localization suggest their potential role in localized endocrine signaling and regulation.