INVESTIGADORES
ELGOYHEN Ana Belen
congresos y reuniones científicas
Título:
Developmental changes in the voltage-gated Ca2+ channels (VGCC) that mediate acetylcholine (ACh) release at the transient efferent-inner hair cell synapse
Autor/es:
KEARNEY G; ZORRILA DE SAN MARTIN J; WEDEMEYER C; ELGOYHEN AB; KATZ E
Reunión:
Congreso; 36th Midwinter Meeting, Association for Research in Otolaryngology; 2014
Resumen:
Since birth until the
onset of hearing (postnatal day (P) 12 in mice), inner hair cells (IHCs) are
innervated by medial olivocochlear fibers. At P9-11, ACh release is supported
by both P/Q and N-type VGCCs and negatively regulated by L-type VGCC, coupled
to BK channel activation. We previously reported that at P5-7, P/Q- but not
N-type VGCC partially support ACh release and that blocking BK channel
increases the quantum content (m) (Zorrilla de San Mart¨ªn et al., ARO Abstracts
2012). Our goal is now to determine which other type/s of VGCCs mediate ACh
release at this (P5-7) and earlier stages (P3) and whether BK channels and
L-type VGCCs are functionally coupled.
Postsynaptic responses were monitored in whole-cell voltage-clamped IHCs
while electrically stimulating the efferent fibers in P3 and P5-7 mouse
cochleas. At P5-7, SNX-482, the R-type VGCC antagonist, significantly reduced m (control: 0.84¡À0.01; 500 nM SNX:
0.37¡À0.05, n=2; p<0.05). Surprisingly, both the L-type VGCC antagonist
(Nifedipine) and the agonist (Bay-K) enhanced m (%increment = 161¡À21, 3 mM Nife; 294¡À26, 10 mM
Bay-K). This suggested that at this earlier stage, Ca2+ entry
through L-type VGCC might be both supporting release and activating BK
channels. However, occlusion experiments showed that Ibtx had no effect on m after incubation with L-type VGCC
modulators (m = 0.56¡À0.18 control;
1.64¡À0.86 Nife; 1.85¡À1.02 Nife+Ibtx; n=2, p>0.50
and m = 1.10¡À0.42 control; 2.67¡À0.80
Bay-K; 2.23¡À0.16 Bay-K+Ibtx; n=3; p
>0.2). Moreover, nifedipine had no effect on m after blocking both P/Q-type VGCC and BK channels with 200 nM
¦Ø-AgaIVA and 100 nM Ibtx, respectively (m
= 0.91¡À0.25 ¦Ø-Aga+Ibtx and 1.08¡À0.29 for ¦Ø-Aga+Ibtx+Nife; p>0.5, n=3). Besides, Bay-K had no effect on m after blocking BK channels (control: 0.95, Ibtx: 2.7, Ibtx+Bay-K: 2.8).
Altogether, these results show that L-type VGCCs do not support but negatively modulate
release by activating BK channels. Preliminary experiments show that at P3,
¦Ø-AgaIVA, does not affect m (control:
054¡À0.06; ¦Ø-AgaIVA 0.62¡À0.25; n=2; p>0.5),
suggesting that P/Q-type VGCCs do not support release at this stage. At P3,
Bay-K significantly enhanced m by
382¡À120%; n=2), suggesting that L-type VGCCs might be also be modulating
release at this stage. We conclude that at P5-7, both P/Q and R-type VGGC
support ACh release and that Ca2+ entry through L-type VGCCs,
negatively modulate this process by activating BK channels.