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Electrogenic Na(+)/Ca(2+) exchange. A novel amplification step in squid olfactory transduction.

Danaceau JP, Lucero MT - J. Gen. Physiol. (2000)

Bottom Line: To directly asses the effects of increasing [Ca(2+)](i) in perforated-patched squid ORNs, we applied 10 mM caffeine to release Ca(2+) from internal stores.We found that electrogenic Na(+)/Ca(2+) exchange was responsible for approximately 26% of the total current associated with glutamate-induced odor responses.Although Na(+)/Ca(2+) exchangers are known to be present in ORNs from numerous species, this is the first work to demonstrate amplifying contributions of the exchanger current to odor transduction.

View Article: PubMed Central - PubMed

Affiliation: Interdepartmental Program in Neuroscience, School of Medicine, Salt Lake City, UT 84108, USA.

ABSTRACT
Olfactory receptor neurons (ORNs) from the squid, Lolliguncula brevis, respond to the odors l-glutamate or dopamine with increases in internal Ca(2+) concentrations ([Ca(2+)](i)). To directly asses the effects of increasing [Ca(2+)](i) in perforated-patched squid ORNs, we applied 10 mM caffeine to release Ca(2+) from internal stores. We observed an inward current response to caffeine. Monovalent cation replacement of Na(+) from the external bath solution completely and selectively inhibited the caffeine-induced response, and ruled out the possibility of a Ca(2+)-dependent nonselective cation current. The strict dependence on internal Ca(2+) and external Na(+) indicated that the inward current was due to an electrogenic Na(+)/Ca(2+) exchanger. Block of the caffeine-induced current by an inhibitor of Na(+)/Ca(2+) exchange (50-100 microM 2',4'-dichlorobenzamil) and reversibility of the exchanger current, further confirmed its presence. We tested whether Na(+)/Ca(2+) exchange contributed to odor responses by applying the aquatic odor l-glutamate in the presence and absence of 2', 4'-dichlorobenzamil. We found that electrogenic Na(+)/Ca(2+) exchange was responsible for approximately 26% of the total current associated with glutamate-induced odor responses. Although Na(+)/Ca(2+) exchangers are known to be present in ORNs from numerous species, this is the first work to demonstrate amplifying contributions of the exchanger current to odor transduction.

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Na+/Ca2+ exchange currents are reversible in squid ORNs. (A) Current traces from a squid ORN under whole-cell voltage-clamp conditions. The solid line above the current trace indicates the switch from ASW to Tris+ ASW. +, 20-mV hyperpolarization. External and internal sodium concentrations are noted above and below the solid line in millimoles per liter−1. Solutions: ASW ext (Tris+ ASW)/Na+ gluconate int with 4 mM Mg2+ and 2 mM ATP. (B) A different ORN exposed to the same protocol as in A, but with Li+ gluconate internal solution to eliminate reverse Na+/Ca2+ exchange. The solid line indicates the switch from ASW to Tris+ ASW and back again. Numbers indicate external and internal sodium concentrations in millimoles per liter−1. +, 20-mV hyperpolarizations. Solutions: ASW ext (Tris+ ASW)/Li+ gluconate int; Rs = 19 MΩ, equilibration time = 5 min). (C) Mean percent decrease in the steady state inward current measured during the switch to Tris ASW under the conditions in A (Na+ int) and B (Li+ int). *The mean current reduction in Li+ gluconate is significantly smaller than Na+ gluconate int P < 0.05. Error bars indicate SD; n = numbers of cells.
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Figure 4: Na+/Ca2+ exchange currents are reversible in squid ORNs. (A) Current traces from a squid ORN under whole-cell voltage-clamp conditions. The solid line above the current trace indicates the switch from ASW to Tris+ ASW. +, 20-mV hyperpolarization. External and internal sodium concentrations are noted above and below the solid line in millimoles per liter−1. Solutions: ASW ext (Tris+ ASW)/Na+ gluconate int with 4 mM Mg2+ and 2 mM ATP. (B) A different ORN exposed to the same protocol as in A, but with Li+ gluconate internal solution to eliminate reverse Na+/Ca2+ exchange. The solid line indicates the switch from ASW to Tris+ ASW and back again. Numbers indicate external and internal sodium concentrations in millimoles per liter−1. +, 20-mV hyperpolarizations. Solutions: ASW ext (Tris+ ASW)/Li+ gluconate int; Rs = 19 MΩ, equilibration time = 5 min). (C) Mean percent decrease in the steady state inward current measured during the switch to Tris ASW under the conditions in A (Na+ int) and B (Li+ int). *The mean current reduction in Li+ gluconate is significantly smaller than Na+ gluconate int P < 0.05. Error bars indicate SD; n = numbers of cells.

Mentions: Voltage-clamp and current-clamp data were acquired with an Axon Instruments 200A patch clamp amplifier (TL-1-125 Interface; Axon Instruments, Inc.) and a 486, 33 MHz computer using either PClamp 5.6 or Axotape 2.0.2. The data in Fig. 1, Fig. 2, and Fig. 3 B were sampled at 250 Hz and filtered off line with a digital eight-pole lowpass Bessel filter (Clampfit 8; Axon Instruments, Inc.) using a −3 dB filter cutoff frequency of 35 Hz. The data in Fig. 3 A, 5, and 6 were sampled at 500 Hz and digitally filtered off line with a −3 dB filter cutoff frequency of 50 Hz. The data in Fig. 4 were acquired at 10 kHz and filtered on line with a low pass four-pole Bessel filter cutoff at 5 kHz. Data for capacitance and series resistance (Rs) measurements were sampled at 100 kHz and also filtered on line at 10 kHz (data not shown). The Rs of nystatin-perforated patched cells averaged 39 ± 13 M (SD; n = 18). Approximately 60–75% of Rs was electrically compensated. The voltage errors associated with the remaining Rs as well as liquid junction potentials were corrected for off line as described in Danaceau and Lucero 1998. All averages are presented as the mean ± SD (n = number of cells).


Electrogenic Na(+)/Ca(2+) exchange. A novel amplification step in squid olfactory transduction.

Danaceau JP, Lucero MT - J. Gen. Physiol. (2000)

Na+/Ca2+ exchange currents are reversible in squid ORNs. (A) Current traces from a squid ORN under whole-cell voltage-clamp conditions. The solid line above the current trace indicates the switch from ASW to Tris+ ASW. +, 20-mV hyperpolarization. External and internal sodium concentrations are noted above and below the solid line in millimoles per liter−1. Solutions: ASW ext (Tris+ ASW)/Na+ gluconate int with 4 mM Mg2+ and 2 mM ATP. (B) A different ORN exposed to the same protocol as in A, but with Li+ gluconate internal solution to eliminate reverse Na+/Ca2+ exchange. The solid line indicates the switch from ASW to Tris+ ASW and back again. Numbers indicate external and internal sodium concentrations in millimoles per liter−1. +, 20-mV hyperpolarizations. Solutions: ASW ext (Tris+ ASW)/Li+ gluconate int; Rs = 19 MΩ, equilibration time = 5 min). (C) Mean percent decrease in the steady state inward current measured during the switch to Tris ASW under the conditions in A (Na+ int) and B (Li+ int). *The mean current reduction in Li+ gluconate is significantly smaller than Na+ gluconate int P < 0.05. Error bars indicate SD; n = numbers of cells.
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Figure 4: Na+/Ca2+ exchange currents are reversible in squid ORNs. (A) Current traces from a squid ORN under whole-cell voltage-clamp conditions. The solid line above the current trace indicates the switch from ASW to Tris+ ASW. +, 20-mV hyperpolarization. External and internal sodium concentrations are noted above and below the solid line in millimoles per liter−1. Solutions: ASW ext (Tris+ ASW)/Na+ gluconate int with 4 mM Mg2+ and 2 mM ATP. (B) A different ORN exposed to the same protocol as in A, but with Li+ gluconate internal solution to eliminate reverse Na+/Ca2+ exchange. The solid line indicates the switch from ASW to Tris+ ASW and back again. Numbers indicate external and internal sodium concentrations in millimoles per liter−1. +, 20-mV hyperpolarizations. Solutions: ASW ext (Tris+ ASW)/Li+ gluconate int; Rs = 19 MΩ, equilibration time = 5 min). (C) Mean percent decrease in the steady state inward current measured during the switch to Tris ASW under the conditions in A (Na+ int) and B (Li+ int). *The mean current reduction in Li+ gluconate is significantly smaller than Na+ gluconate int P < 0.05. Error bars indicate SD; n = numbers of cells.
Mentions: Voltage-clamp and current-clamp data were acquired with an Axon Instruments 200A patch clamp amplifier (TL-1-125 Interface; Axon Instruments, Inc.) and a 486, 33 MHz computer using either PClamp 5.6 or Axotape 2.0.2. The data in Fig. 1, Fig. 2, and Fig. 3 B were sampled at 250 Hz and filtered off line with a digital eight-pole lowpass Bessel filter (Clampfit 8; Axon Instruments, Inc.) using a −3 dB filter cutoff frequency of 35 Hz. The data in Fig. 3 A, 5, and 6 were sampled at 500 Hz and digitally filtered off line with a −3 dB filter cutoff frequency of 50 Hz. The data in Fig. 4 were acquired at 10 kHz and filtered on line with a low pass four-pole Bessel filter cutoff at 5 kHz. Data for capacitance and series resistance (Rs) measurements were sampled at 100 kHz and also filtered on line at 10 kHz (data not shown). The Rs of nystatin-perforated patched cells averaged 39 ± 13 M (SD; n = 18). Approximately 60–75% of Rs was electrically compensated. The voltage errors associated with the remaining Rs as well as liquid junction potentials were corrected for off line as described in Danaceau and Lucero 1998. All averages are presented as the mean ± SD (n = number of cells).

Bottom Line: To directly asses the effects of increasing [Ca(2+)](i) in perforated-patched squid ORNs, we applied 10 mM caffeine to release Ca(2+) from internal stores.We found that electrogenic Na(+)/Ca(2+) exchange was responsible for approximately 26% of the total current associated with glutamate-induced odor responses.Although Na(+)/Ca(2+) exchangers are known to be present in ORNs from numerous species, this is the first work to demonstrate amplifying contributions of the exchanger current to odor transduction.

View Article: PubMed Central - PubMed

Affiliation: Interdepartmental Program in Neuroscience, School of Medicine, Salt Lake City, UT 84108, USA.

ABSTRACT
Olfactory receptor neurons (ORNs) from the squid, Lolliguncula brevis, respond to the odors l-glutamate or dopamine with increases in internal Ca(2+) concentrations ([Ca(2+)](i)). To directly asses the effects of increasing [Ca(2+)](i) in perforated-patched squid ORNs, we applied 10 mM caffeine to release Ca(2+) from internal stores. We observed an inward current response to caffeine. Monovalent cation replacement of Na(+) from the external bath solution completely and selectively inhibited the caffeine-induced response, and ruled out the possibility of a Ca(2+)-dependent nonselective cation current. The strict dependence on internal Ca(2+) and external Na(+) indicated that the inward current was due to an electrogenic Na(+)/Ca(2+) exchanger. Block of the caffeine-induced current by an inhibitor of Na(+)/Ca(2+) exchange (50-100 microM 2',4'-dichlorobenzamil) and reversibility of the exchanger current, further confirmed its presence. We tested whether Na(+)/Ca(2+) exchange contributed to odor responses by applying the aquatic odor l-glutamate in the presence and absence of 2', 4'-dichlorobenzamil. We found that electrogenic Na(+)/Ca(2+) exchange was responsible for approximately 26% of the total current associated with glutamate-induced odor responses. Although Na(+)/Ca(2+) exchangers are known to be present in ORNs from numerous species, this is the first work to demonstrate amplifying contributions of the exchanger current to odor transduction.

Show MeSH
Related in: MedlinePlus