Added after amiloride, Phe had no statistically significant effects on Cl? secretion by HTGM or CF-HTGM (Fig. 6A). By contrast, MCh, ATP, and Iso all stimulated Isc of HTGM with Kd values of ~3 �� 10?7 M and maximal responses by 10?5 M. The effectiveness of secretagogues on increasing Isc across HTGM SB203580 PKB was ATP > MCh Iso > Phe (Fig. 6A). Responses of CFTGM to MCh, ATP, and Phe were not significantly different from those of HTGM (Fig. 6A). By contrast, the response of CFTGM to Iso was significantly reduced to ~20% that of HTGM (Fig. 6A). Fig. 6. Chloride and mucin secretion by HTGM and cystic fibrosis (CF) human tracheobronchial gland mucous (CFTGM) cells in response to adrenergic, cholinergic, and purinergic agonists (10?5 M). A: chloride secretion measured in Ussing chambers under short-circuit .

.. Effects of secretagogues on mucin secretion. The basal rate of mucin secretion, measured in the period immediately before addition of secretagogue (i.e., the third 30-min collection period), was not statistically different between HTGM (68 �� 11 ng?cm?2?h?1, n = 56) and CFTGM (100 �� 16 ng?cm?2?h?1, n = 14). However, for both HTGM and CFTGM, control (untreated) tissues showed a significant decline in secretion over time, such that during the fourth collection period (the period during which drugs were added to test tissues), mucin secretion from HTGM was 77 �� 3% of that in the third period, whereas in CFTGM cells it was 79 �� 7%. Therefore, as described in methods, the secretory responses of test tissues were corrected for this time-dependent decline.

Preliminary experiments showed that the responses of CFTGM to mediators had Kd values much the same as for normal cells. Therefore, responses were tested in detail for 10?5 M only. Figure 6B shows that responses to MCh and ATP were reduced in CFTGM cells, but that the responses to Iso and PE were unaltered. Effects of Cl? channel blockade. In HTGM cells, the nonspecific blocker of Cl? channels, DPAC (1 mM), significantly reduced the response in Isc to all stimulatory secretagogues. In eight pairs of tissues, Iso-induced chloride secretion was reduced from 9.3 �� 2.0 to 0. 8 �� 0.2 ��A/cm2 (P < 0.05), ATP-induced chloride secretion was reduced from 15.0 �� 1.8 to 1. 5 �� 0.5 ��A/cm2 (P < 0.05), and MCh-induced secretion was reduced from 10.5 �� 1. 4 to 4.3 �� 1.3 ��A/cm2 (P < 0.05).

The response to Phe was not significantly different from zero, either with or Brefeldin_A without the blocker. Despite its actions on the Isc responses to mediators, DPAC had little or no effect on mediator-induced mucin secretion. Thus, in eight sheets from three tracheas, the secretory index for Iso was 155 �� 12 in the absence of blocker and 144 �� 15 in the presence. Corresponding values for Phe were 123 �� 14 and 133 �� 12; for ATP, 226 �� 15 and 177 �� 40; and for MCh, 133 �� 22 and 83 �� 8, respectively. DISCUSSION Airway gland mucous secretions are abnormally viscous in CF (33).