No abstract

Cystic fibrosis is caused by mutations in CFTR, the cystic fibrosis transmembrane conductance regulator gene. Disruption of CFTR-mediated anion conductance results in defective fluid and electrolyte movement in the epithelial cells of organs such as the pancreas, airways and sweat glands, but the function of CFTR in salivary glands is unclear. Salivary gland acinar cells produce an isotonic, plasma-like fluid, which is subsequently modified by the ducts to produce a hypotonic, NaCl-depleted final saliva. In the present study we investigated whether submandibular salivary glands (SMGs) in F508 mice (Cftr F/ F ) display ion transport defects characteristic of cystic fibrosis in other tissues. Immunolocalization and whole-cell recordings demonstrated that Cftr and the epithelial Na + (ENaC) channels are co-expressed in the apical membrane of submandibular duct cells, consistent with the significantly higher saliva [NaCl] observed in vivo in Cftr F/ F mice. In contrast, Cftr and ENaC channels were not detected in acinar cells, nor was saliva production affected in Cftr F/ F mice, implying that Cftr contributes little to the fluid secretion process in the mouse SMG. To identify the source of the NaCl absorption defect in Cftr F/ F mice, saliva was collected from ex vivo perfused SMGs. Cftr F/ F glands secreted saliva with significantly increased [NaCl]. Moreover, pharmacological inhibition of either Cftr or ENaC in the ex vivo SMGs mimicked the Cftr F/ F phenotype. In summary, our results demonstrate that NaCl absorption requires and is likely to be mediated by functionally dependent Cftr and ENaC channels localized to the apical membranes of mouse salivary gland duct cells.

No abstract

No abstract

Salivary glands express multiple isoforms of P2X and P2Y nucleotide receptors, but their in vivo physiological roles are unclear. P2 receptor agonists induced salivation in an ex vivo submandibular gland preparation. The nucleotide selectivity sequence of the secretion response was BzATP Ͼ Ͼ ATP > ADP Ͼ Ͼ UTP, and removal of external Ca 2؉ dramatically suppressed the initial ATP-induced fluid secretion (ϳ85%). Together, these results suggested that P2X receptors are the major purinergic receptor subfamily involved in the fluid secretion process. Mice with targeted disruption of the P2X 7 gene were used to evaluate the role of the P2X 7 receptor in nucleotide-evoked fluid secretion. P2X 7 receptor protein and BzATPactivated inward cation currents were absent, and importantly, purinergic receptor agonist-stimulated salivation was suppressed by more than 70% in submandibular glands from P2X 7 -null mice. Consistent with these observations, the ATP-induced increases in [Ca 2؉ ] i were nearly abolished in P2X 7 ؊/؊ submandibular acinar and duct cells. ATP appeared to also act through the P2X 7 receptor to inhibit muscarinic-induced fluid secretion. These results demonstrate that the ATP-sensitive P2X 7 receptor regulates fluid secretion in the mouse submandibular gland.Salivation is a Ca 2ϩ -dependent process (1, 2) primarily associated with the neurotransmitters norepinephrine and acetylcholine, release of which stimulates ␣-adrenergic and muscarinic receptors, respectively. Both types of receptors are coupled to G proteins that activate phospholipase C␤ (PLC␤) during salivary gland stimulation. PLC␤ activation cleaves phosphatidylinositol 1,4-bisphosphate resulting in diacylglycerol and inositol 1,4,5-trisphosphate (InsP 3 ) production. Activation of Ca 2ϩ -selective InsP 3 receptor channels localized to the endoplasmic reticulum of salivary acinar cells increases the intracellular free calcium concentration ([Ca 2ϩ ] i ). 4 Depletion of the endoplasmic reticulum Ca 2ϩ pool triggers extracellular Ca 2ϩ influx and a sustained elevation in [Ca 2ϩ ] i . This increase in [Ca 2ϩ ] i activates Ca 2ϩ -dependent K ϩ and Cl Ϫ channels promoting Cl Ϫ secretion across the apical membrane and a lumen negative, electrochemical gradient that supports Na ϩ efflux into the lumen. The accumulation of NaCl creates an osmotic gradient which drives water movement into the lumen, thus generating isotonic primary saliva. This primary fluid is then modified by the ductal system, which reabsorbs NaCl and secretes KHCO 3 producing a final saliva that is hypotonic (1, 2).Salivation also has a non-cholinergic, non-adrenergic component, the origin of which is unclear (3). In addition to muscarinic and ␣-adrenergic receptors, salivary acinar cells express other receptors that are coupled to an increase in [Ca 2ϩ ] i such as purinergic P2 and substance P receptors. Like muscarinic and ␣-adrenergic receptors, P2 receptor activation leads to a sustained increase in [Ca 2ϩ ] i in salivary acinar cells (4). In contrast, substance P receptor ac...

We have recently shown that the IK1 and maxi-K channels in parotid salivary gland acinar cells are encoded by the K Ca 3.1 and K Ca 1.1 genes, respectively, and in vivo stimulated parotid secretion is severely reduced in double-null mice. The current study tested whether submandibular acinar cell function also relies on these channels. We found that the K + currents in submandibular acinar cells have the biophysical and pharmacological footprints of IK1 and maxi-K channels and their molecular identities were confirmed by the loss of these currents in K Ca 3.1-and K Ca 1.1-null mice. Unexpectedly, the pilocarpine-stimulated in vivo fluid secretion from submandibular glands was essentially normal in double-null mice. This result and the possibility of side-effects of pilocarpine on the nervous system, led us to develop an ex vivo fluid secretion assay. Mammals express three major paired salivary glands, the parotid, submandibular and sublingual. The morphology, histology and saliva composition are unique for each gland type (Young & Van Lennep, 1977). These differences are most evident in the secretory endpieces, also known as acini, the cells which secrete the water and most of the protein found in saliva. In rodents, the parotid acinar cells are serous, while submandibular acinar cells are seromucous. In contrast, the secretory endpieces of sublingual glands are mixed in nature, containing both mucous and serous demilune cells. The composition of the saliva collected from individual glands varies according V. G. Romanenko and T. Nakamoto contributed equally to this work. This paper has online supplemental material.to the acinar cell type. Parotid glands produce a watery fluid and sublingual glands secrete a thick, ropey saliva due to the exocytosis of heavily glycosylated mucins, whereas submandibular saliva has an intermediate consistency.The mechanism of fluid secretion is generally studied at two levels: (i) the cellular level gives detailed information under well controlled conditions but is an over-simplified model and (ii) the in vivo level provides access to whole organ function, but the interpretation of results is complicated by systemic effects. In the latter case, saliva secretion is often induced by cholinergic agonists such as pilocarpine. It is well established that pilocarpine can stimulate saliva secretion through glandular muscarinic receptors. However, systemic administration of a cholinergic agonist also activates receptors in the brain and peripheral nervous

No abstract

No abstract