The salivary glands of hard ticks consist of three types (type I, II, and III) of acini according to their functions and location. The type II and III acini play critical roles in tick salivation, which is likely controlled by a variety of neuropeptides or neurotransmitter via interaction with their receptor, G protein-coupled receptors (GPCRs). Orchestration of dopamine receptor (D1) and invertebrate specific D1-like dopamine receptor (InvD1L) located in type II and III acini precisely control tick salivary secretion via collection of primary saliva in the lumen and expulsion of collected saliva, respectively. The two dopamine receptors (D1 and InvD1L) in Haemaphysalis longicornis were identified as 1278 bp (426 aa) and 1362 bp (454 aa) in length, respectively. Both dopamine receptors were functionally analyzed through Ca2+ and cAMP assay using the heterologous expression system. The transcripts of D1 and InvD1L were profiled from synganglion and salivary glands of female ticks (unfed, 3/18/60/96 post blood meal and replete). D1 and InvD1L were significantly upregulated in the early phase of blood feeding from female H. longicornis. Salivary secretion induced by dopamine was significantly reduced by RNAi of D1 and InvD1L. Interestingly, RNAi of two dopamine receptors induced a significantly longer period of blood feeding in female ticks, which were significantly lighter after feeding than control. Taken together, it was suggested that D1 and InvD1L play critical roles in early and late phase of tick blood feeding for feeding capability.

The effects of adenosine triphosphate(ATP) on salivary glands have been recognized since 1982. The presence of purinergic recepetors(P2Rs) that mediate the effects of ATP in various tissues, including parotid and submandibular salivary gland, has been supported by the cloning of receptor cDNAs and the expression of the receptor proteins. P2Rs have many subtypes, and the activation of these receptor subtypes increase intracellular Cα²+, a key ion in the regulation of the secretion in the salivary gland. The apical pores of taste buds in circumvallate and foliate papillae are surrounded by the saliva from von Ebner salivary gland(vEG). Thus, it is important how the secretion of vEG is controlled. This study was designed to elucidate the roles of P2Rs on salivary secretion of vEG. Male Sprague-Dawley rats (about 200 g) were used for this experiment. vEG-rich tissues were obtained from dissecting 500-1,000μm thick posterior tongue slices under stereomicroscope view. P2Rs mRNA in vEG acinar cells were identified with RT-PCR. To observe the change in intracellular Cα²+ activity, we employed Cα²+-ion specific fluorescence analysis with fura-2. Single acinar cells and cell clusters were isolated by a sequential trypsin/collagenase treatment and were loaded with 10 μM fura -2 AM for 60 minutes at room temperature. Several agonists and antagonists were used to test a receptor specificity. RT-PCR revealed that the mRNAs of P2X₄, P2Y₁, P2Y₂ and P2Y₃ are expressed in vEG acinar cells. The intracellular calcium activity was increased in response to 10 μM ATP, a P2Rs agonist, and 2-MeSATP, a P2Y₄ and P2Y₂R agonist. However, 300 μM αβ-MeATP, a P2X₁ and P2X₃R agonist, did not elicit the response. The responses elicited by 10 μM ATP and UTP, a P2Y₂R agonists, were maintained when extracellular calcium was removed. 10 μM suramin, a P2XR antagonist, and reactive blue 2, a P2YR antagonist, partially blocked ATP-induced response. However, when extracellular calciums were removed, suramin did not abolish the responses elicited by ATP. These results suggest that P2Rs play an important role in salivary secretion of vEG acinar cells and the effects of ATP on vEG salivary secretion may be mediated by P2X₄, P2Y₁, P2Y₂, and/or P2Y₃.