[Show abstract][Hide abstract] ABSTRACT: Previous studies have shown that galvanic stimulation of semicircular canal organs can modulate their afferent discharge. However, it has not been resolved whether this modulation derived from direct stimulation of hair cells, afferent nerve fibers, some combination of the two, or some as yet unknown path. This problem is addressed in the present study. Experiments were designed first to determine the gross current path necessary for the DC current to modulate afferent firing. These led to the conclusion that the current path had to flow between endolymph and perilymph across the neuroepithelium. Next, the various components in this established path were considered: the afferents, the hair cells, between the hair cells, or some combination of the three. These experiments led to the conclusion that the current pathway was across the hair cells causing transmitter release and thus affecting afferent activity.
[Show abstract][Hide abstract] ABSTRACT: Hair cells in mechanosensory systems transduce mechanical stimuli into biological signals to be presented to and analyzed by the brain. Vestibular hair cells transduce stimuli primarily associated with the organism's orientation and motion in space. When examined superficially it may appear that the hair cells act as passive transducers whereby mechanical stimulation of their hair bundle results in transmitter release at their afferent synapses. In fact, hair cell functions are more complicated, and the mechanical signals are heavily processed even before being encoded in afferent nerve activity. Hair cells are different from one another in morphology, biophysics, transmitter and transmitter receptor complements, not only across different organs (as one might expect), but even in the same organ. This review focuses on hair cell morpho-physiological properties, ionic conductances, neurotransmitters/modulators and their receptors, second messengers and effectors. Special features of hair cell neurotransmission, as the synaptic body and the presence of autoreceptors and local circuits, are also discussed, as is the possibility of a differential modulation of hair cell transmitter release in the resting and mechanically-stimulated states.
Progress in Neurobiology 03/1998; 54(2):193-247. · 9.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Studies of K+ conductances in hair cells report that big-conductance Ca(2+)-dependent K+ (BK) channels carry parts of the outwardly rectifying currents. Lin et al. (1995) suggested that in guinea pig outer hair cells (OHCs) a portion of these currents is carried via a voltage-dependent and Ca(2+)-independent K+ channel. The present study tests the hypothesis that there are two separable current components of the outwardly rectifying currents by using patch clamp methods in OHCs to characterize the voltage dependence and sensitivity of the outwardly rectifying currents to channel blockers. Lowering of external Ca2+ caused no change in the currents while charybdotoxin (ChTx; 100 nM), a BK K+ channel blocker, and Cd2+ (200 microM), and L-type calcium channel blocker, abolished about 50% of the currents. Both ChTx and Cd2+ caused a depolarizing shift in the half-activation voltage paralleled by a decrease in the voltage sensitivity. 4-Aminopyridine (4-AP, 0.01 mM), an A-type and delayed rectifier type channel blocker, abolished about 50% of the currents and caused a hyperpolarizing shift in the half-activation voltage together with an increase in the voltage sensitivity. The outwardly rectifying currents were more sensitive to block by 4-AP at membrane voltages around 40 mV compared to voltages around -20 mV. The differences in the current characteristics may be due to two separate channel types, one of which is similar to the delayed rectifier type channels while the other may be similar to the BK Ca(2+)-dependent K+ channels. In addition, the largest outwardly rectifying currents were present in long OHCs with the smallest present in short OHCs.
Hearing Research 04/1997; 105(1-2):146-58. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The type of K+ channel involved in the acetylcholine (ACh) evoked response (Ksub; sub stands for suberyldicholine) in guinea pig outer hair cells (OHCs) is still uncertain. The present study tests the hypotheses that Ksub is one of the following: a big conductance Ca(2+)-dependent K+ channel (BK), a small conductance Ca(2+)-dependent K+ channel (SK), a KA type of K+ channel, or a Kn type of K+ channel. Patch-clamp technique in the whole-cell mode was used to record from single guinea pig OHCs. ACh (100 microM) was applied to voltage-clamped OHCs and the ACh-induced currents (IACh) were measured. Charybdotoxin (100 nM) had no effect on IACh, while apamin (1 microM) blocked more than 90% of IACh. Lowering the external Ca2+ concentration caused a hyperpolarizing shift of the IACh monitored as a function of the prepulse voltage. Increasing internal Mg2+ (Mgi2+) concentration caused a reduction in the outward IACh without affecting the inward IACh. The Ksub channel was found to be permeable to Cs+. In Cs+ solutions, IACh was 45% of the IACh in K+ solutions. The block of IACh by apamin, the dependence on extracellular Ca2+, the incomplete block of IACh by Cs+, and the ACh-induced Cs+ currents favor the hypothesis that Ksub belongs to the SK type of channels. An ionotropic/nicotinic nature of the ACh mechanism of action is favored. It is suggested that, in vivo, the amplitude of the ACh-induced hyperpolarization may depend on the Ca2+/Mg2+ ratio inside and outside the cell.
Hearing Research 12/1996; 101(1-2):149-72. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The properties of the ACh (acetylcholine) response in guinea pig outer hair cells (OHCs) are not well understood. It has been shown that the response to ACh involves the activation of a Ca2+ dependent K+ selective conductance (referred to as Ksub where sub stands for suberyldicholine). In the present study, we examined the voltage dependence, the time dependence, and the desensitization of the ACh response. In addition, we examined the K+ selectivity of K(sub). These properties are important for aiding in the determination of the type of K+ channels activated by ACh. Patch-clamp technique in the whole-cell mode was used to record from single OHCs isolated from adult pigmented guinea pigs. ACh (100 microM) was applied to the voltage-clamped OHCs and the ACh induced currents (IACh) were measured. A voltage dependence of the ACh response was found with the ACh induced currents decaying monoexponentially at potentials positive to -30 mV. The decay of the ACh induced currents was faster soon after establishing the whole-cell mode of recording when compared to the decay of the currents some time later. This effect, referred to as the time dependence, was different from the desensitization of the response upon prolonged application of ACh. The desensitization of the ACh induced currents was about 50% after 2 min of continuous application of 100 microM ACh. The examined characteristics of the ACh response in guinea pig OHCs indicate a voltage and time dependence of the response and strong K+ selectivity of the Ksub.
Hearing Research 12/1996; 101(1-2):132-48. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In this article the evidence concerning the nature of the acetylcholine (ACh) receptors on hair cells is reviewed. A schematic organization of these receptors is offered, based on the evidence as follows. (1) There are two kinds of ACh receptors on hair cells: muscarinic-like and nicotinic-like. (2) The nicotinic-like receptor mediates a hyperpolarizing response to ACh and a consequent reduction in afferent firing. (3) The muscarinic-like receptors mediate both a depolarization and a hyperpolarization of hair cells. (4) The hyperpolarization results in a reduction in afferent firing and (5) the depolarization results in an increase in afferent firing.
Hearing Research 10/1996; 98(1-2):1-8. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Biochemical and pharmacological evidence supports a role for nitric oxide (NO) in the cochlea. In the present experiments, we tested sodium nitroprusside (SNP), an NO donor, applied by intracochlear perfusions on sound-evoked responses of the cochlea (CM, cochlear microphonic; SP, summating potential; EP, endocochlear potential; CAP, compound action potential) and in vitro on outer hair cell (OHC) voltage-induced length changes and current responses. In vivo application of SNP in increasing concentrations (10, 33, 100, 330 and 1000 microM) reduced all sound-evoked responses starting at about 300 microM. The responses continued to decline after a postdrug wash. At 1 mM SNP decreased EP slowly (approximately 80 min) whereas at 10 mM it reduced EP more rapidly (approximately 20 min). Ferricyanide (1 mM) and S-nitroso-N-acetylpenicillamine (SNAP; 1 mM) had no effect on sound-evoked cochlear potentials. Ferricyanide (1 mM and 10 mM) and ferrocyanide (10 mM) had no effect on EP. In vitro, SNP (10 mM) significantly reduced both OHC voltage-induced length changes and whole-cell outward currents. Results suggest that SNP, possibly acting by released NO, influences cochlear function through effects at the stria vascularis and at the OHCs.
Hearing Research 08/1995; 87(1-2):1-8. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ca2+ channel currents and their modulation by adenosine 5'-triphosphate (ATP) in acutely isolated guinea pig outer hair cells (OHCs) were investigated using the whole-cell patch-clamp technique. The current-voltage (I-V) relation of OHCs indicated that the Ca2+ channel opened near -30 mV, and the current reached a maximum at +10 and 0 mV in 20 mM Ca2+ and Ba2+ external solutions, respectively. BayK 8644 (BayK, 2 microM) caused a 3.5-fold increase in peak Ca2+ currents and shifted the I-V curves toward more negative potentials. These results suggest that the majority of Ca2+ channels in OHCs have L-type characteristics. The effects of ATP on Ca2+ channels of OHCs were heterogenous. ATP (100 microM) decreased Ca2+ channel currents by 31.7 +/- 5.6% at 0 mV and shifted Ca2+ tail activation curves toward more depolarized potentials in some cells (N = 6). By contrast, in others, ATP enhanced the currents by 43.5 +/- 12.5% at +10 mV (N = 6). In the presence of BayK, however, ATP-induced inhibition or enhancement of Ca2+ channel currents was attenuated. In addition, 100 microM ATP produced little effect on Ca2+ channel currents in another subpopulation of cells (N = 12). This heterogenous neuromodulation of Ca2+ channel currents by ATP may reflect a functional diversity among OHCs.
Hearing Research 07/1995; 86(1-2):25-33. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Previous studies have shown that low concentrations of exogenous ATP, added to the perilymphatic fluid, could modify the bioelectrical activity of the isolated semicircular canal of the frog (Rana pipiens). To test the hypothesis that ATP is endogenously present and active in the perilymphatic fluid, the influence of two ATP-purinoceptor antagonists, Reactive Blue 2 and suramin, and of the enzyme, nucleotide pyrophosphatase, were examined. When applied by perilymphatic bath substitution, the three compounds reduced, in a dose-dependent manner, the firing of the afferent fibers monitored in the absence of mechanically-applied stimulation. The response of the afferent fibers, recorded when the sensory cells were mechanically inhibited, was also reduced. No modification of the response of the excitatory phase of the mechanical stimulus was observed in the presence of the two antagonists. In contrast, the signal was significantly reduced by the enzyme. None of the three compounds exhibited an influence on the transepithelial potential, or its variation in response to mechanical stimulation. The ATP-induced modification of the firing rate of the afferent fibers, monitored in the absence of mechanical stimulation, was reduced in the presence of the three drugs. No influence of Reactive Blue 2 and suramin was observed on the increase of the spontaneous firing induced by carbachol. In contrast, the effect of carbachol was decreased by nucleotide pyrophosphatase. The excitatory influence of glutamate on the spontaneous firing was not modified by Reactive Blue 2, while it was slightly increased by suramin and nucleotide pyrophosphatase.(ABSTRACT TRUNCATED AT 250 WORDS)
[Show abstract][Hide abstract] ABSTRACT: Acetylcholine (ACh), the major neurotransmitter released by efferent nerve fibers in the cochlea, has been shown to activate a Ca(2+)-dependent K+ conductance in outer hair cells (OHCs). Previously we reported that this ACh operated conductance is permeable to Cs+. The purpose of the present study was to characterize further this Cs(+)-permeable channel and its dependency on Ca2+ using isolated OHCs and the patch clamp technique in the whole cell configuration. The changes in the ACh response were examined when Cs+, Ba2+, Cd2+, N-methyl-D-glucamine (NMG+) and tetraethylammonium (TEA+) were placed in the external or internal solutions. Cs+ substituted for K+ in carrying the ACh-evoked Ca(2+)-dependent K+ current. When NMG+/TEA+ was substituted for internal K+ ACh-evoked an inward and an outward current, and Cs+ substituted for external K+ blocked the outward but not the inward current evoked by ACh suggesting it was carried by K+. In the NMG+/TEA+ condition, when the cell was held at different Vh values for an extended period of time, the ACh-induced K+ current rectified. In Ba2+ (3 mM) with zero Ca2+ ACh failed to induce any detectable current and the ACh response slowly recovered from the Ba2+ block, suggesting a block at an intracellular site. Cd2+ (1 mM) readily and reversibly blocked ACh-induced currents even when carried by Cs+. This data suggests that ACh opens a channel selective for K+, conductive to Cs+ and dependent on Ca2+.
Hearing Research 01/1995; 81(1-2):119-29. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In the present study, the influence of extracellular ATP and ATP agonists in the physiology of the vestibular organs was examined, using the in vitro model of the isolated semicircular canal of the frog (Rana pipiens). The firing activity of the afferent nerve, the d.c. nerve potential and the transepithelial potential were measured in the absence and presence of mechanical stimulation of the sensory epithelium. Administration of ATP into the perilymphatic compartment, from 10(-12) to 10(-3) M, increased the firing rate of the afferent fibers recorded in the absence of mechanical stimulation. Recordings of the d.c. nerve potential indicated that the afferent fibers were hyperpolarized. The presence of the purine also modified the transepithelial potential. During mechanical stimulation of the sensory epithelium, both the evoked afferent firing and the evoked variation of the d.c. nerve potential were reduced in the presence of ATP. However, ATP did not effect the evoked modulation of the transepithelial potential, evoked by the mechanical stimulation. Administration of the P2x purinoceptor agonists, alpha, beta-methylene-ATP and beta, gamma-methylene-ATP, at concentrations between 10(-12) and 10(-3) M, did not significantly modify the different bioelectrical activities investigated. In contrast, 2-methylthio-ATP, a P2y purinoceptor agonist, more potent and efficacious than ATP in its effect on the spontaneous firing. Concurrently, no modification of the d.c. nerve potential, the transepithelial potential and their variation during mechanical stimulation was observed. In opposition to the ATP effect, the total amplitude of the evoked firing was increased in the presence of 2-methylthio-ATP. These data suggest that extracellular ATP, present in the perilymphatic compartment, may act as a neuromodulator in the vestibular physiology. The effects of the purine appear to be mediated by the activation of a P2y subtype of purinoceptor. The absence of an effect of ATP and 2-methylthio-ATP on the evoked variation of the transepithelial potential suggest that the purine did not affect the processes responsible for the generation of the receptor potential but more likely modified the mechanisms involved in the release of the neurotransmitter from the hair cells and/or acted on the afferent endings.
[Show abstract][Hide abstract] ABSTRACT: In the present study, the influence of extracellular ATP and ATP agonists in the physiology of the vestibular organs was examined, using thein vitro model of the isolated semicircular canal of the frog (Rana pipiens). The firing activity of the afferent nerve, the d.c. nerve potential and the transepithelial potential were measured in the absence and presence of mechanical stimulation of the sensory epithelium. Administration of ATP into the perilymphatic compartment, from 10−12 to 10−3M, increased the firing rate of the afferent fibers recorded in the absence of mechanical stimulation. Recordings of the d.c. nerve potential indicated that the afferent fibers were hyperpolarized. The presence of the purine also modified the transepithelial potential. During mechanical stimulation of the sensory epithelium, both the evoked afferent firing and the evoked variation of the d.c. nerve potential were reduced in the presence of ATP. However, ATP did not effect the evoked modulation of the transepithelial potential, evoked by the mechanical stimulation. Administration of the P2x purinoceptor agonists, α,β-methylene-ATP and β, γ-methylene-ATP, at concentrations between 10−12 and 10−3 M, did not significantly modify the different bioelectrical activities investigated. In contrast, 2-methylthio-ATP, a P2y purinoceptor agonist, more potent and efficacious than ATP in its effect on the spontaneous firing. Concurrently, no modification of the d.c. nerve potential, the transepithelial potential and their variation during mechanical stimulation was observed. In opposition to the ATP effect, the total amplitude of the evoked firing was increased in the presence of 2-methylthio-ATP.These data suggest that extracellular ATP, present in the perilymphatic compartment, may act as a neuromodulator in the vestibular physiology. The effects of the purine appear to be mediated by the activation of a P2y subtype of purinoceptor. The absence of an effect of ATP and 2-methylthio-ATP on the evoked variation of the transepithelial potential suggest that the purine did not affect the processes responsible for the generation of the receptor potential but more likely modified the mechanisms involved in the release of the neurotransmitter from the hair cells and/or acted on the afferent endings.
[Show abstract][Hide abstract] ABSTRACT: Stimulation of the efferent nerves to the vestibular organs of the frog's inner ear produces either facilitation or inhibition of afferent firing. Similarly, application of acetylcholine (ACH), the major transmitter of the efferents, can produce both facilitation and/or inhibition as previously reported [Guth et al. (1986) Acta Otolaryngol. 102, 194-204; Norris et al. (1988) Hear. Res. 32, 197-206]. The firing rates of afferent neurons of the semicircular canal (SCC) using multiunit recordings are generally facilitated by ACH. Conversely, the firing rates of afferent units innervating the saccule are generally inhibited by ACH. This latter inhibition is antagonized by strychnine more potently than by curare, which is more potent than atropine. When inhibition is antagonized by strychnine or curare an underlying facilitation is revealed. The inhibition of saccular afferents by ACH shows desensitization requiring about 20 min to recover. The ACH-induced inhibition is mimicked by nicotine at very high concentrations but not by dimethyl phenylpiperazinium or cytisine. The fact that multiunit afferent firing from the SCC is generally facilitated while that from the saccule is generally inhibited by ACH suggests a different distribution of ACH receptors and receptor types (i.e. muscarinic or nicotinic and their subtypes) in the two organs and demonstrates the usefulness of recording from multiple units simultaneously. The difference in distribution of ACH receptors may be important for understanding the physiology of vestibular efferents.
Hearing Research 06/1994; 75(1-2):225-32. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Acetylcholine (ACh) is the major neurotransmitter released from the efferent fibers in the cochlea onto the outer hair cells (OHCs). The type of ACh receptor on OHCs and the events subsequent to receptor activation are unclear. Therefore we studied the effect of agonists and antagonists of the ACh receptor on isolated OHCs from the guinea pig. OHCs were recorded from in whole cell voltage and current clamp configuration. ACh induced an increase in outward K+ current (IACh) which hyperpolarized the OHCs. No desensitization to ACh application was observed. Cs+ replaced K+ in carrying the IACh. The IACh is Ca(2+)-dependent, time and voltage sensitive, and different from the IKCa induced by depolarization of the membrane potential. When tested at 100 microM, several agonists also induced outward current responses (acetylcholine > suberyldicholine > or = carbachol > DMPP) whereas nicotine, cytisine and muscarine did not. The IACh response to 10 microM ACh was blocked by low concentrations of traditional and non-traditional-nicotinic antagonists (strychnine > curare > bicuculline > alpha-bungarotoxin > thimethaphan) and by higher concentrations of muscarinic antagonists (atropine > 4-DAMP > AF-DX 116 > pirenzepine). Pharmacologically, the ACh receptor on OHCs is nicotinic.
Hearing Research 05/1994; 74(1-2):135-47. · 2.54 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The morphological characteristics of isolated guinea-pig outer hair cells (OHCs) and frog semicircular canal hair cells (SCCHCs) were reviewed. Active and passive electrophysiological properties of OHCs and SCCHCs were compared utilizing the whole cell variant of the patch clamp technique. Results lead to the conclusion that both hair cell types are similarly viable. SCCHCs were dominated by inactivating outward conductances and were strongly inwardly rectified. In contrast, OHCs demonstrated little inactivation or rectification. The differences in electrophysiological properties between the two cell types may reflect the separate functions these cells have in the ear.
Comparative Biochemistry and Physiology Part A: Physiology. 01/1994;
[Show abstract][Hide abstract] ABSTRACT: In developing the technique of selective chemical vestibulectomy, the destructive effect of streptomycin on the hair cells of the inner ear was well established. However, in both animal and human studies, a rapid onset of the drug's vestibular inhibition was observed in addition to a more slowly developing long-term destructive effect. In previous laboratory studies, streptomycin had been given only by a systemic route, and only the chronic long-term effects had been observed. In this study, an isolated semicircular canal was prepared and streptomycin was placed into the perilymph bathing the canal. The spontaneous and evoked afferent nerve activity were recorded prior to streptomycin application, during application, and after a washout period. During streptomycin application, the activity of the semicircular canal was reversibly inhibited in a dose dependent manner. After a sufficient washout period (5-10 min), the preparation had completely recovered from the drug's effect. Thus, there are two phases in the vestibular inhibition by streptomycin: an early reversible phase that subsequently can transform into a later irreversible phase.
The American journal of otology 08/1993; 14(4):373-9.
[Show abstract][Hide abstract] ABSTRACT: Pharmacologists are among the most recent basic medical scientists to apply themselves to the auditory and vestibular systems. They bring to the subject a fresh perspective and, uniquely, the ability to control biological processes through drugs. Interest in the auditory and vestibular systems among pharmacologists has focussed on: the drug-sensitive processes (primarily but not exclusively neuro-transmission); toxicology, and pharmacotherapy. Pharmacology holds the promise of shedding further light on auditory and vestibular function and ameliorating auditory and vestibular dysfunction.
[Show abstract][Hide abstract] ABSTRACT: 1. A-type outward currents were studied in sensory hair cells isolated from the semicircular canals (SCC) of the leopard frog (Rana pipiens) with whole-cell voltage- and current-clamping techniques. 2. There appear to be two classes of A-type outward-conducting potassium channels based on steady-state, kinetic, pharmacological parameters, and reversal potential. 3. The two classes of A-type currents differ in their steady-state inactivation properties as well as in the kinetics of inactivation. The steady-state inactivation properties are such that a significant portion of the fast channels are available from near the resting potential. 4. The inactivating channels studied do not appear to be calcium dependent. 5. The A-channels in hair cells appear to subserve functions that are analogous to IA functions in neurons, that is, modulating spike latency and Q (the oscillatory damping function). The A-currents appear to temporally limit the hair cell voltage response to a current injection.
Journal of Neurophysiology 12/1992; 68(5):1642-53. · 3.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: It has been generally accepted that even in the absence of mechanical stimulation of the transductional elements, a resting depolarizing current exists which is ultimately responsible for the spontaneous release of neurotransmitter. Movement of the transductional elements modulates this resting current and thereby the evoked release of neurotransmitter occurs. Recent data from our laboratory and others have led us to question whether the relationship between spontaneous and evoked neurotransmitter release is as simple as stated. Indeed, a variety of experimental manipulations appear to influence the two modes of release differently. Examination of our results and the results of others has led us to four hypotheses: 1. the two modes of neurotransmitter release are processed differently by the hair cells; 2. cyclic AMP is involved in spontaneous but not evoked neurotransmitter release; 3. there is a positive feedback step involving an excitatory amino acid and its receptor on the hair cell in evoked neurotransmitter release and; 4. different pools of calcium are involved according to the mode of release. Accordingly, there may be several biochemical steps between the transductional movement of the stereocilia at the apex of the hair cells and the ultimate release of the neurotransmitter at the base of these cells. Some of these biochemical steps are different depending on whether the mode of release is spontaneous or evoked. These biochemical steps may amplify or at least interact with the biophysical processes previously described in the hair cells.
Hearing Research 12/1991; 56(1-2):69-78. · 2.54 Impact Factor