POLIVINIL-ACRÍLICAS BASE ACUOSA
3. RESULTADOS Y DISCUSIÓN
3.1 CONDICIONES ADECUADAS PARA LA OBTENCIÓN Y CARACTERIZACIÓN DE EMULSIONES.
Inward rectifier channels have a major role in maintaining the resting
membrane potential of RBL-2H3 cells. Ba^^ is a known inhibitor of the IR channel in RBL-2H3 cells. Nevertheless, though it can completely inhibit IR channels at lOOpM, it had little effect on stimulated secretion in RBL-2H3 cells, at that concentration and even up to ImM. However, Ba^^ inhibited antigen-induced secretion with an IC50 of about
2mM. The effects of Ba^^ on the RBL-2H3 cell responses will be discussed later in section 7.4.
CsCl also blocks the IR channel in RBL-2H3 cells. It inhibits this current completely at ImM, but more than lOmM was needed to have a significant effect on antigen-stimulated 8-hexosaminidase release. Cs"^ blocks IR channels ( at 2mM ) [209,
211] and OR channels (at 20mM) in RBL-2H3 cells [218]. In view of the inhibition of IR current by 2mM Cs^ and the lack of effect of Cs"^ on antigen-induced secretion these results provided further evidence that an IR current is not involved in RBL-2H3 cell activation.
TEA has been reported to inhibit IR channels in RBL-2H3 cells at a
concentration between 10-15mM [208, 209]. However, as already mentioned, it did not inhibit antigen-induced degranulation. Since charybdotoxin, but not TEA, inhibited antigen-stimulated secretion in RBL-2H3 cells, it is unlikely that the possible channels
are similar to BKca channels. However, the data presented so far suggest that the possible channels resemble IKca channels.
Amiodarone, an inhibitor of the IR channel in the heart [258], was tested and it did not have any inhibitory effect on antigen-mediated activation of RBL-2H3 cells.
Taken together, these results indicate that IR channels do not play an important role in the antigen-induced signal transduction mechanisms of RBL-2H3 cells.
3.4.2.10 ATP-sensitive channel blocker
Glibenclamide, a specific blocker of ATP-sensitive channels in B-cells of the pancreas and skeletal muscle, even at concentration of lOpM, had no significant effect on
antigen-induced B-hexosaminidase release. Its reported IC50 for ATP-sensitive channel
blocking is 4-27nM. Therefore, Katp channels do not seem to be involved in antigen- activated secretion in RBL-2H3 cells.
3.4.2.11 K a t p channel openers
Cromakalim, a non-specific channel opener, and diazoxide, a selective ATP- sensitive channel opener, did not induce any significant inhibitory effect on antigen-
induced B-hexosaminidase release. Thus, it seems that the possible channel involved in
antigen-stimulated RBL-2H3 cells is not sensitive to cromakalim or diazoxide.
To summarise, the inhibitory effect of quinidine, charybdotoxin, cetiedil and the UCL compounds may suggest that the potassium channels possibly involved in antigen- induced activation of RBL-2H3 cells may resemble the IKca channels in red blood cells.
Table 3.7: The K channel modulators tested, their ICso on channels and the measured IC20 & IC50 on antigen- stimulated P-hexosaminidase release in RBL-2H3.
1 Quinidine different channels Na^ channels Voltage-dependent Ca^* channel 15-400pM SR '= 10p M ER^=15pM SR =30pM E R =45pM 2 4 6 , 2 5 9 , 2 6 0 2 BaClj IR K' channels OR K' channels lOOpM 2-20m M S R = lm M E R =1.5m M SR =2m M ER=6m M 2 0 8 3 CsCl IR K channel OR K" channels 0.5m M 20mM > lOmM > 20m M 2 0 9 , 2 1 1 , 2 1 8
4 Glibenclam ide ATP-sensitive K' channels
4-27nM 60pM N D 2 6 1 , 2 6 2
5 Apamin SKca channels l-3nM N S at 1 uM N S at IpM 2 6 3 , 2 6 4
6 U K 6 6 9 1 4 Delayed rectifier & IR K* channels
0 .3 -lp M N S at lOpM N S at lOpM 257 Nitrendipine OR K^ channels 1.3pM SR =0.4pM SR =1.3pM 210 Nicardipine L-type Ca^* channels 0.7p M E R = > 1 0 p M E R = N D 245 8 Charybdotoxm BKca channels
IK(\. channels
l-30nM 30nM
35nM 90nM 2 6 5 , 2 6 6 , 2 6 7
9 ΠA Diiferent K channels 0.3-10m M N S at 20m M N S at 20m M 2 4 6 , 2 4 7 , 2 4 8
10 Amiodarone IR K^ channels 20pM N S at 50pM N S at SOpM 2 5 8
11 Cetiedil IKca channels 25-150p M 25pM 90pM 254 12 Ib'I'X BKca l-3nM N S a t lOOnM N S at lOOnM 2 6 8
13 M gTX Kvi 3 < InM N S at lOOnM N S at lOOnM 176 14 Dequalinium SKca channels 1-1,5pM N S at 50pM N S at 50pM 2 5 2 , 2 6 4
15 Clotrimazole IKca channels l-24n M 25pM 70pM 251 16 Propafenone different K' channel Na^ channel 5pM N S a t lOpM N S a t lOpM 2 6 0 , 2 6 9 17 9- Ammoacridme
SKca and Katp 70pM N S at 60pM N S at 60 u M 198, 246 18 4-A P different K" channels 5pM -3m M 2mM 6m M 2 4 9 , 2 5 0
19 Cromakalim different K^ channels 0 .1 -lp M N S at 50pM N S at 50p M 2 7 0
20 Diazoxide ATP-sensiti\ e K* channel
20pM N S a t lOOpM N S at lOOpM 2 6 2
*-Serotonin release +-P-hexosaminldase (enzvme) release J-Not tested NS- No Significant effect ND- not determined * T h ese references are related to the effect of i C channel modulators and their IC sn in other tissues.