scholarly journals Fast Adaptation in Mouse Olfactory Sensory Neurons Does Not Require the Activity of Phosphodiesterase

2006 ◽  
Vol 128 (2) ◽  
pp. 171-184 ◽  
Author(s):  
Anna Boccaccio ◽  
Laura Lagostena ◽  
Volker Hagen ◽  
Anna Menini
Keyword(s):  
Sensory Neurons ◽  
Molecular Mechanisms ◽  
Flash Photolysis ◽  
Physiological Role ◽  
Reversal Potential ◽  
Cell Voltage ◽  
Olfactory Sensory Neurons ◽  
Cng Channel ◽  
Ion Substitution ◽  
Fast Adaptation

Vertebrate olfactory sensory neurons rapidly adapt to repetitive odorant stimuli. Previous studies have shown that the principal molecular mechanisms for odorant adaptation take place after the odorant-induced production of cAMP, and that one important mechanism is the negative feedback modulation by Ca2+-calmodulin (Ca2+-CaM) of the cyclic nucleotide-gated (CNG) channel. However, the physiological role of the Ca2+-dependent activity of phosphodiesterase (PDE) in adaptation has not been investigated yet. We used the whole-cell voltage-clamp technique to record currents in mouse olfactory sensory neurons elicited by photorelease of 8-Br-cAMP, an analogue of cAMP commonly used as a hydrolysis-resistant compound and known to be a potent agonist of the olfactory CNG channel. We measured currents in response to repetitive photoreleases of cAMP or of 8-Br-cAMP and we observed similar adaptation in response to the second stimulus. Control experiments were conducted in the presence of the PDE inhibitor IBMX, confirming that an increase in PDE activity was not involved in the response decrease. Since the total current activated by 8-Br-cAMP, as well as that physiologically induced by odorants, is composed not only of current carried by Na+ and Ca2+ through CNG channels, but also by a Ca2+-activated Cl− current, we performed control experiments in which the reversal potential of Cl− was set, by ion substitution, at the same value of the holding potential, −50 mV. Adaptation was measured also in these conditions of diminished Ca2+-activated Cl− current. Furthermore, by producing repetitive increases of ciliary's Ca2+ with flash photolysis of caged Ca2+, we showed that Ca2+-activated Cl− channels do not adapt and that there is no Cl− depletion in the cilia. All together, these results indicate that the activity of ciliary PDE is not required for fast adaptation to repetitive stimuli in mouse olfactory sensory neurons.

Temporal Development of Cyclic Nucleotide-Gated and Ca2+-Activated Cl− Currents in Isolated Mouse Olfactory Sensory Neurons

2007 ◽  
Vol 98 (1) ◽  
pp. 153-160 ◽  
Author(s):  
Anna Boccaccio ◽  
Anna Menini
Keyword(s):  
Sensory Neurons ◽  
Flash Photolysis ◽  
Olfactory Receptors ◽  
Cell Voltage ◽  
Equilibrium Potential ◽  
Olfactory Sensory Neurons ◽  
Second Phase ◽  
Current Response ◽  
Intact Mouse ◽  
Secondary Current

A Ca2+-activated Cl− current constitutes a large part of the transduction current in olfactory sensory neurons. The binding of odorants to olfactory receptors in the cilia produces an increase in cAMP concentration; Ca2+ enters into the cilia through CNG channels and activates a Cl− current. In intact mouse olfactory sensory neurons little is known about the kinetics of the Ca2+-activated Cl− current. Here, we directly activated CNG channels by flash photolysis of caged cAMP or 8-Br-cAMP and measured the current response with the whole cell voltage-clamp technique in mouse neurons. We measured multiphasic currents in the rising phase of the response at −50 mV. The current rising phase became monophasic in the absence of extracellular Ca2+, at +50 mV, or when most of the intracellular Cl− was replaced by gluconate to shift the equilibrium potential for Cl− to −50 mV. These results show that the second phase of the current in mouse intact neurons is attributed to a Cl− current activated by Ca2+, similarly to previous results on isolated frog cilia. The percentage of the total saturating current carried by Cl− was estimated in two ways: 1) by measuring the maximum secondary current and 2) by blocking the Cl− channel with niflumic acid. We estimated that in the presence of 1 mM extracellular Ca2+ and in symmetrical Cl− concentrations the Cl− component can constitute up to 90% of the total current response. These data show how to unravel the CNG and Ca2+-activated Cl− component of the current rising phase.

scholarly journals The voltage dependence of the TMEM16B/anoctamin2 calcium-activated chloride channel is modified by mutations in the first putative intracellular loop

2012 ◽  
Vol 139 (4) ◽  
pp. 285-294 ◽  
Author(s):  
Valentina Cenedese ◽  
Giulia Betto ◽  
Fulvio Celsi ◽  
O. Lijo Cherian ◽  
Simone Pifferi ◽  
...  
Keyword(s):  
Structure Function ◽  
Sensory Neurons ◽  
Molecular Mechanisms ◽  
Voltage Dependence ◽  
Cell Voltage ◽  
Site Directed Mutagenesis ◽  
Initial Structure ◽  
Intracellular Loop ◽  
Retinal Photoreceptors ◽  
Cl Channels

Ca2+-activated Cl− channels (CaCCs) are involved in several physiological processes. Recently, TMEM16A/anoctamin1 and TMEM16B/anoctamin2 have been shown to function as CaCCs, but very little information is available on the structure–function relations of these channels. TMEM16B is expressed in the cilia of olfactory sensory neurons, in microvilli of vomeronasal sensory neurons, and in the synaptic terminals of retinal photoreceptors. Here, we have performed the first site-directed mutagenesis study on TMEM16B to understand the molecular mechanisms of voltage and Ca2+ dependence. We have mutated amino acids in the first putative intracellular loop and measured the properties of the wild-type and mutant TMEM16B channels expressed in HEK 293T cells using the whole cell voltage-clamp technique in the presence of various intracellular Ca2+ concentrations. We mutated E367 into glutamine or deleted the five consecutive glutamates 386EEEEE390 and 399EYE401. The EYE deletion did not significantly modify the apparent Ca2+ dependence nor the voltage dependence of channel activation. E367Q and deletion of the five glutamates did not greatly affect the apparent Ca2+ affinity but modified the voltage dependence, shifting the conductance–voltage relations toward more positive voltages. These findings indicate that glutamates E367 and 386EEEEE390 in the first intracellular putative loop play an important role in the voltage dependence of TMEM16B, thus providing an initial structure–function study for this channel.

scholarly journals Calcium-activated chloride channels in the apical region of mouse vomeronasal sensory neurons

2012 ◽  
Vol 140 (1) ◽  
pp. 3-15 ◽  
Author(s):  
Michele Dibattista ◽  
Asma Amjad ◽  
Devendra Kumar Maurya ◽  
Claudia Sagheddu ◽  
Giorgia Montani ◽  
...  
Keyword(s):  
Sensory Neurons ◽  
Chloride Channels ◽  
Calcium Concentration ◽  
Transient Receptor Potential ◽  
Flash Photolysis ◽  
Physiological Role ◽  
Disulfonic Acid ◽  
Apical Region ◽  
Caged Calcium ◽  
Vomeronasal Sensory Neurons

The rodent vomeronasal organ plays a crucial role in several social behaviors. Detection of pheromones or other emitted signaling molecules occurs in the dendritic microvilli of vomeronasal sensory neurons, where the binding of molecules to vomeronasal receptors leads to the influx of sodium and calcium ions mainly through the transient receptor potential canonical 2 (TRPC2) channel. To investigate the physiological role played by the increase in intracellular calcium concentration in the apical region of these neurons, we produced localized, rapid, and reproducible increases in calcium concentration with flash photolysis of caged calcium and measured calcium-activated currents with the whole cell voltage-clamp technique. On average, a large inward calcium-activated current of −261 pA was measured at −50 mV, rising with a time constant of 13 ms. Ion substitution experiments showed that this current is anion selective. Moreover, the chloride channel blockers niflumic acid and 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid partially inhibited the calcium-activated current. These results directly demonstrate that a large chloride current can be activated by calcium in the apical region of mouse vomeronasal sensory neurons. Furthermore, we showed by immunohistochemistry that the calcium-activated chloride channels TMEM16A/anoctamin1 and TMEM16B/anoctamin2 are present in the apical layer of the vomeronasal epithelium, where they largely colocalize with the TRPC2 transduction channel. Immunocytochemistry on isolated vomeronasal sensory neurons showed that TMEM16A and TMEM16B coexpress in the neuronal microvilli. Therefore, we conclude that microvilli of mouse vomeronasal sensory neurons have a high density of calcium-activated chloride channels that may play an important role in vomeronasal transduction.

Odorant Responses of Dual Polarity Are Mediated by cAMP in Mouse Olfactory Sensory Neurons

2004 ◽  
Vol 92 (3) ◽  
pp. 1312-1319 ◽  
Author(s):  
Rona Delay ◽  
Diego Restrepo
Keyword(s):  
Sensory Neurons ◽  
Olfactory Sensory Neurons ◽  
Olfactory Responses ◽  
Outward Currents ◽  
Cng Channel ◽  
Patch Configuration ◽  
Excitatory Responses ◽  
Dual Polarity ◽  
Camp Levels ◽  
A Current

Some olfactory sensory neurons (OSNs) respond to odors with hyperpolarization. Although transduction for excitatory responses is mediated by opening of a cyclic nucleotide-gated (CNG) channel, there is controversy on the mechanism underlying inhibitory responses. We find that mouse OSNs respond to odorants by either depolarizing or hyperpolarizing responses in loose-patch measurements. In the perforated-patch configuration, OSNs not only responded with a current consistent with CNG channel-mediated excitation but also displayed enhancement of outward currents, consistent with inhibitory responses. Increasing cAMP levels pharmacologically elicited excitatory or inhibitory responses in different OSNs. In addition, OSNs from mice defective for the CNGA2 subunit of the CNG channel displayed neither excitatory nor inhibitory responses. Thus CNG channels mediate inhibitory olfactory responses.

scholarly journals PACS‐1 Mediates Phosphorylation‐Dependent Ciliary Trafficking of the CNG Channel in Olfactory Sensory Neurons

2009 ◽  
Vol 23 (S1) ◽  
Author(s):  
Paul Michael Jenkins ◽  
Lian Zhang ◽  
Gary Thomas ◽  
Jeffrey R. Martens
Keyword(s):  
Sensory Neurons ◽  
Olfactory Sensory Neurons ◽  
Cng Channel

Nonselective cation channels coupled with tachykinin receptors in rat sensory neurons

1995 ◽  
Vol 73 (2) ◽  
pp. 736-742 ◽  
Author(s):  
K. Inoue ◽  
K. Nakazawa ◽  
K. Inoue ◽  
K. Fujimori
Keyword(s):  
Sensory Neurons ◽  
Reversal Potential ◽  
Cell Voltage ◽  
Cation Channels ◽  
Neurokinin A ◽  
Extracellular Solution ◽  
Inward Current ◽  
Nonselective Cation Channels ◽  
Excitatory Neurotransmission ◽  
In Cells

1. Effects of substance P (SP) and other tachykinins on membrane currents were investigated using whole cell voltage clamp in cultured sensory neurons isolated from rat dorsal root ganglia. 2. SP (100 nM) evoked an inward current in two-thirds of the cells at negative potentials. In most of the cells that generated the inward current in response to SP, capsaicin also activated an inward current. The SP-evoked inward current was not observed in cells loaded with 2 mM guanosine 5'-O-(2-thiodiphosphate) (GDP beta S). 3. Neurokinin A (NKA) or neurokinin B (NKB) also activated an inward current. At 100 nM of each agonist, the order was NKB > NKA > SP with respect to activated current amplitude. 4. The tachykinin-activated current was reversed around +10 mV with a standard extracellular solution containing 140 mM NaCl. The reversal potential became more negative when extracellular NaCl was reduced by substituting with sucrose. The inward current was also activated in cells bathed in an extracellular solution containing Cs+, tetraethylammonium (TEA) or N-methyl-D-glucamine (NMDG) as a major cation instead of Na+. The order of permeability, determined from the reversal potential of the current, was Cs+ not equal to Na+ > TEA > NMDG. The amplitude of the inward current activated by NKB was increased when extracellular Na+ was replaced with Cs+, TEA or NMDG. 5. Permeability of Ca2+ was tested using an extracellular solution containing Ca2+ as the only cation (111.8 mM Ca2+ outside). Under this condition, NKB evoked an inward current that reversed around +30 mV. 6. The results suggest that SP and other tachykinins activate nonselective cation channels, which are also permeable to Ca2+, through receptors which are more responsive to NKB than to SP or NKA. The channel activation may serve as a mechanism underlying tachykinin-mediated excitatory neurotransmission in sensory neurons.

Antagonistic Modulation of a Hyperpolarization-Activated Cl– Current in Aplysia Sensory Neurons by SCPB and FMRFamide

2003 ◽  
Vol 90 (2) ◽  
pp. 586-598 ◽  
Author(s):  
Ned Buttner ◽  
Steven A. Siegelbaum
Keyword(s):  
Sensory Neurons ◽  
Reversal Potential ◽  
Outward Current ◽  
Cell Voltage ◽  
High Concentration ◽  
Pipette Solution ◽  
Whole Cell ◽  
Whole Cell Recording ◽  
Type K ◽  
Cell Recording

Whole cell voltage-clamp recordings from Aplysia mechanosensory neurons obtained from the pleural ganglion were used to investigate the actions on membrane currents of the neuropeptides SCPB and FMRFamide. At the start of whole cell recording, SCPB typically evoked an inward current at a holding potential of –40 mV, due to the cAMP-mediated closure of the S-type K+ channel, whereas FMRFamide evoked an outward current, due to the opening of the S-type K+ channels mediated by 12-lipoxygenase metabolites of arachidonic acid. However, after several minutes of whole cell recording with a high concentration of chloride in the whole cell patch pipette solution, the responses to SCPB and FMRF-amide at –40 mV were inverted; SCPB evoked an outward current, whereas FMRFamide and YGGFMRFamide evoked inward currents. Ion substitution experiments and reversal potential measurements revealed that these responses were due to the opposing regulation of a Cl– current, whose magnitude was greatly enhanced by dialysis with the high Cl–-containing pipette solution. SCPB inhibited this Cl– current through production of cAMP and activation of PKA. YGGFMRFamide activated this Cl– current by stimulating a cGMP-activated phosphodiesterase that hydrolyzed cAMP. Thus a cAMP-dependent Cl– current undergoes antagonistic modulation by two neuropeptides in Aplysia sensory neurons.

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