The cardiac hormone atrial natriuretic peptide (ANP) signals via interaction with a plasma membrane receptor which has guanylyl cyclase (GC) activity and is referred to as GC-A. Desensitization of GC-A is thought to represent a physiologically important regulatory mechanism, but the signaling pathways implicated and cell type-specific effects are still poorly understood. Here we demonstrate that sustained exposure to either ANP itself or to the bioactive lipid lysophosphatidic acid (LPA) elicits GC-A desensitization in MA-10 Leydig cells. Both reactions show similar kinetics and evoke equal decreases (by 40%) in GC-A hormone-responsiveness. Homologous (ANP-induced) desensitization, where cyclic GMP is generated as second messenger, is blocked by distinct cyclic AMP-dependent protein kinase (PKA) inhibitors, H 89 and Rp-8-CPT-cAMPS, providing evidence that PKA mediates the reaction. Accordingly, the ANP/cGMP-elicited effects are mimicked by a cAMP analog, 8-Br-cAMP. The LPA-induced (heterologous) desensitization is not blocked by PKA inhibition, indicating a different signaling pathway. LPA, but not ANP, enhances extracellular signal-regulated kinase (ERK) phosphorylation and induces cell rounding together with a dramatic re-organization of actin filaments. Consistent with the identification of LPA receptor (LPA₂ and LPA₃) gene expression, the findings are indicative of LPA receptor-mediated reactions. This study demonstrates for the first time co-existence of homologous and heterologous desensitization of GC-A in the same cell type, reveals that these reactions are mediated by different pathways and identifies a novel crosstalk between phospholipid and natriuretic peptide signaling. The morphoregulatory activities exerted by LPA suggest a crucial role for Leydig cell physiology.