Hair Cell Synapses Research Articles

Review of Hair Cell Synapse Defects in Sensorineural Hearing Impairment

To review new insights into the pathophysiology of sensorineural hearing impairment. Specifically, we address defects of the ribbon synapses between inner hair cells and spiral ganglion neurons that cause auditory synaptopathy. Here, we review original publications on the genetics, animal models, and molecular mechanisms of hair cell ribbon synapses and their dysfunction. Hair cell ribbon synapses are highly specialized to enable indefatigable sound encoding with utmost temporal precision. Their dysfunctions, which we term auditory synaptopathies, impair audibility of sounds to varying degrees but commonly affect neural encoding of acoustic temporal cues essential for speech comprehension. Clinical features of auditory synaptopathies are similar to those accompanying auditory neuropathy, a group of genetic and acquired disorders of spiral ganglion neurons. Genetic auditory synaptopathies include alterations of glutamate loading of synaptic vesicles, synaptic Ca influx or synaptic vesicle turnover. Acquired synaptopathies include noise-induced hearing loss because of excitotoxic synaptic damage and subsequent gradual neural degeneration. Alterations of ribbon synapses likely also contribute to age-related hearing loss.

Quantitative analysis of ribbons, vesicles, and cisterns at the cat inner hair cell synapse: Correlations with spontaneous rate

Cochlear hair cells form ribbon synapses with terminals of the cochlear nerve. To test the hypothesis that one function of the ribbon is to create synaptic vesicles from the cisternal structures that are abundant at the base of hair cells, we analyzed the distribution of vesicles and cisterns around ribbons from serial sections of inner hair cells in the cat, and compared data from low and high spontaneous rate (SR) synapses. Consistent with the hypothesis, we identified a "sphere of influence" of 350 nm around the ribbon, with fewer cisterns and many more synaptic vesicles. Although high- and low-SR ribbons tended to be longer and thinner than high-SR ribbons, the total volume of the two ribbon types was similar. There were almost as many vesicles docked at the active zone as attached to the ribbon. The major SR-related difference was that low-SR ribbons had more synaptic vesicles intimately associated with them. Our data suggest a trend in which low-SR synapses had more vesicles attached to the ribbon (51.3 vs. 42.8), more docked between the ribbon and the membrane (12 vs. 8.2), more docked at the active zone (56.9 vs. 44.2), and more vesicles within the "sphere of influence" (218 vs. 166). These data suggest that the structural differences between high- and low-SR synapses may be more a consequence, than a determinant, of the physiological differences.

Inspiration from Bertram Scharf’s work

In 1990, Bertram Scharf and I discussed about me doing a post doc in his laboratory. The opportunity to work with Bertram did not materialize, but his work in loudness, efferents, and attention has been a continuing inspiration not only for my own research but also for auditory perception, physiology, and audio engineering in general. For example, a Google Scholar search of “Bertram Scharf” on November 8, 2012, produced 1517 citations for his top 10 papers, with 6 on loudness and critical bands, 2 on efferents, and 2 on attention. Here I highlight two recent projects that have been inspired by Scharf’s work. The first project showed significant loudness adaptation in patients with auditory neuropathy, particularly those with otoferlin deficits. This result directly supports Scharf’s proposition that simple loudness adaptation is due to a sensory process, which in this case can be pinned down to transmitter release and replenishment in the hair cell and nerve synapse. The second project extended Scharf’s theoretical work in efferents and attention to improving feedback control in cochlear implant users and tinnitus sufferers. This line of work could improve cochlear implant speech perception in noise and reduce internal gain to alleviate tinnitus.

Neural-scaled entropy as a model of information for speech perception

Neural-Scaled Entropy (NSE) is an objective metric used to quantify “information” available in speech consequent hearing loss, hearing aid signal processing, and distortion from various environmental factors. One pursuit is to use NSE to find optimum hearing aid settings that maximize speech perception. Inspired by the Cochlear-Scaled Entropy model [Stilp et al., J. Acoust. Soc. Am. 2112–2126 (2010)], NSE uses the neural spike output at the inner hair cell synapse of an auditory nerve model [Zilany et al., J. Acoust. Soc. Am. 126, 2390–2412 (2009)]. Probability of spike output from fibers sampled at equidistant places along the model cochlea is computed for short duration time frames. Potential information is estimated by using the Kullback-Liebler Divergence to describe how the pattern of neural firing at each frame differs from preceding frames in an auto-regressive manner. NSE was tested using nonsense syllables from various perceptual studies that included different signal processing schemes and was compared to performance for different vowel-defining parameters, consonant features, and talker gender. NSE has potential to serve as a model predictor of speech perception, and to capture the effects of sensorineural hearing loss beyond simple filter broadening. [Work supported by NIDCD RC1DC010601.]

Single Ca2+ channels and exocytosis at sensory synapses

Hair cell synapses in the ear and photoreceptor synapses in the eye are the first synapses in the auditory and visual system. These specialized synapses transmit a large amount of sensory information in a fast and efficient manner. Moreover, both small and large signals with widely variable kinetics must be quickly encoded and reliably transmitted to allow an animal to rapidly monitor and react to its environment. Here we briefly review some aspects of these primary synapses, which are characterized by a synaptic ribbon in their active zones of transmitter release. We propose that these synapses are themselves highly specialized for the task at hand. Photoreceptor and bipolar cell ribbon synapses in the retina appear to have versatile properties that permit both tonic and phasic transmitter release. This allows them to transmit changes of both luminance and contrast within a visual field at different ambient light levels. By contrast, hair cell ribbon synapses are specialized for a highly synchronous form of multivesicular release that may be critical for phase locking to low-frequency sound-evoked signals at both low and high sound intensities. The microarchitecture of a hair cell synapse may be such that the opening of a single Ca(2+) channel evokes the simultaneous exocytosis of multiple synaptic vesicles. Thus, the differing demands of sensory encoding in the eye and ear generate diverse designs and capabilities for their ribbon synapses.

ATP-gated ion channels mediate adaptation to elevated sound levels

The sense of hearing is remarkable for its auditory dynamic range, which spans more than 10(12) in acoustic intensity. The mechanisms that enable the cochlea to transduce high sound levels without damage are of key interest, particularly with regard to the broad impact of industrial, military, and recreational auditory overstimulation on hearing disability. We show that ATP-gated ion channels assembled from P2X2 receptor subunits in the cochlea are necessary for the development of temporary threshold shift (TTS), evident in auditory brainstem response recordings as sound levels rise. In mice null for the P2RX2 gene (encoding the P2X2 receptor subunit), sustained 85-dB noise failed to elicit the TTS that wild-type (WT) mice developed. ATP released from the tissues of the cochlear partition with elevation of sound levels likely activates the broadly distributed P2X2 receptors on epithelial cells lining the endolymphatic compartment. This purinergic signaling is supported by significantly greater noise-induced suppression of distortion product otoacoustic emissions derived from outer hair cell transduction and decreased suprathreshold auditory brainstem response input/output gain in WT mice compared with P2RX2-null mice. At higher sound levels (≥95 dB), additional processes dominated TTS, and P2RX2-null mice were more vulnerable than WT mice to permanent hearing loss due to hair cell synapse disruption. P2RX2-null mice lacked ATP-gated conductance across the cochlear partition, including loss of ATP-gated inward current in hair cells. These data indicate that a significant component of TTS represents P2X2 receptor-dependent purinergic hearing adaptation that underpins the upper physiological range of hearing.

Electrocochleography in Meniere's disease: recovery from adaptation of the auditory action potential

Conclusions: High prevalence rate of an enhanced summating potential (SP)/action potential (AP) ratio (≥0.35) is considered to result largely from the imbalances of Ca2+ homeostasis in the endolymphatic space. An increase in Ca2+ is thought to augment the SP. Furthermore, a greater inflow of Ca2+ into inner hair cells is supposed to influence the disturbed release of neurotransmitter, which may induce pathological functioning of inner hair cell synapses. We speculate that such abnormal synaptic functioning is responsible for the development of abnormally increased AP recovery from adaptation (iR). Objectives: This study aimed to record the AP recovery from adaptation (AP recovery) and to investigate its clinical significance in association with other electrocochleography (ECochG) findings in Meniere's disease. Methods: ECochG was performed transtympanically in 27 patients with Meniere's disease. AP recovery was measured by a paired-click stimulation paradigm as a function of interclick intervals from 5 to 100 ms. Results: High prevalence of SP/AP ratio was observed in 27 patients. iR, suggesting abnormal functioning of inner hair cell synapse, was in many cases combined with an enhanced SP/AP ratio.

Glutamate Transporters EAAT4 and EAAT5 Are Expressed in Vestibular Hair Cells and Calyx Endings

Glutamate is the neurotransmitter released from hair cells. Its clearance from the synaptic cleft can shape neurotransmission and prevent excitotoxicity. This may be particularly important in the inner ear and in other sensory organs where there is a continually high rate of neurotransmitter release. In the case of most cochlear and type II vestibular hair cells, clearance involves the diffusion of glutamate to supporting cells, where it is taken up by EAAT1 (GLAST), a glutamate transporter. A similar mechanism cannot work in vestibular type I hair cells as the presence of calyx endings separates supporting cells from hair-cell synapses. Because of this arrangement, it has been conjectured that a glutamate transporter must be present in the type I hair cell, the calyx ending, or both. Using whole-cell patch-clamp recordings, we demonstrate that a glutamate-activated anion current, attributable to a high-affinity glutamate transporter and blocked by DL-TBOA, is expressed in type I, but not in type II hair cells. Molecular investigations reveal that EAAT4 and EAAT5, two glutamate transporters that could underlie the anion current, are expressed in both type I and type II hair cells and in calyx endings. EAAT4 has been thought to be expressed almost exclusively in the cerebellum and EAAT5 in the retina. Our results show that these two transporters have a wider distribution in mice. This is the first demonstration of the presence of transporters in hair cells and provides one of the few examples of EAATs in presynaptic elements.

Lack of Brain-Derived Neurotrophic Factor Hampers Inner Hair Cell Synapse Physiology, But Protects against Noise-Induced Hearing Loss

The precision of sound information transmitted to the brain depends on the transfer characteristics of the inner hair cell (IHC) ribbon synapse and its multiple contacting auditory fibers. We found that brain derived neurotrophic factor (BDNF) differentially influences IHC characteristics in the intact and injured cochlea. Using conditional knock-out mice (BDNF(Pax2) KO) we found that resting membrane potentials, membrane capacitance and resting linear leak conductance of adult BDNF(Pax2) KO IHCs showed a normal maturation. Likewise, in BDNF(Pax2) KO membrane capacitance (ΔC(m)) as a function of inward calcium current (I(Ca)) follows the linear relationship typical for normal adult IHCs. In contrast the maximal ΔC(m), but not the maximal size of the calcium current, was significantly reduced by 45% in basal but not in apical cochlear turns in BDNF(Pax2) KO IHCs. Maximal ΔC(m) correlated with a loss of IHC ribbons in these cochlear turns and a reduced activity of the auditory nerve (auditory brainstem response wave I). Remarkably, a noise-induced loss of IHC ribbons, followed by reduced activity of the auditory nerve and reduced centrally generated wave II and III observed in control mice, was prevented in equally noise-exposed BDNF(Pax2) KO mice. Data suggest that BDNF expressed in the cochlea is essential for maintenance of adult IHC transmitter release sites and that BDNF upholds opposing afferents in high-frequency turns and scales them down following noise exposure.

Phylogenetic differences in calcium permeability of the auditory hair cell cholinergic nicotinic receptor

The α9 and α10 cholinergic nicotinic receptor subunits assemble to form the receptor that mediates efferent inhibition of hair cell function within the auditory sensory organ, a mechanism thought to modulate the dynamic range of hearing. In contrast to all nicotinic receptors, which serve excitatory neurotransmission, the activation of α9α10 produces hyperpolarization of hair cells. An evolutionary analysis has shown that the α10 subunit exhibits signatures of positive selection only along the mammalian lineage, strongly suggesting the acquisition of a unique function. To establish whether mammalian α9α10 receptors have acquired distinct functional properties as a consequence of this evolutionary pressure, we compared the properties of rat and chicken recombinant and native α9α10 receptors. Our main finding in the present work is that, in contrast to the high (pCa(2+)/pMonovalents ∼10) Ca(2+) permeability reported for rat α9α10 receptors, recombinant and native chicken α9α10 receptors have a much lower permeability (∼2) to this cation, comparable to that of neuronal α4β2 receptors. Moreover, we show that, in contrast to α10, α7 as well as α4 and β2 nicotinic subunits are under purifying selection in vertebrates, consistent with the conserved Ca(2+) permeability reported across species. These results have important consequences for the activation of signaling cascades that lead to hyperpolarization of hair cells after α9α10 gating at the cholinergic-hair cell synapse. In addition, they suggest that high Ca(2+) permeability of the α9α10 cholinergic nicotinic receptor might have evolved together with other features that have given the mammalian ear an expanded high-frequency sensitivity.

Role for a Novel Usher Protein Complex in Hair Cell Synaptic Maturation

The molecular mechanisms underlying hair cell synaptic maturation are not well understood. Cadherin-23 (CDH23), protocadherin-15 (PCDH15) and the very large G-protein coupled receptor 1 (VLGR1) have been implicated in the development of cochlear hair cell stereocilia, while clarin-1 has been suggested to also play a role in synaptogenesis. Mutations in CDH23, PCDH15, VLGR1 and clarin-1 cause Usher syndrome, characterized by congenital deafness, vestibular dysfunction and retinitis pigmentosa. Here we show developmental expression of these Usher proteins in afferent spiral ganglion neurons and hair cell synapses. We identify a novel synaptic Usher complex comprised of clarin-1 and specific isoforms of CDH23, PCDH15 and VLGR1. To establish the in vivo relevance of this complex, we performed morphological and quantitative analysis of the neuronal fibers and their synapses in the Clrn1−/− mouse, which was generated by incomplete deletion of the gene. These mice showed a delay in neuronal/synaptic maturation by both immunostaining and electron microscopy. Analysis of the ribbon synapses in Ames waltzerav3J mice also suggests a delay in hair cell synaptogenesis. Collectively, these results show that, in addition to the well documented role for Usher proteins in stereocilia development, Usher protein complexes comprised of specific protein isoforms likely function in synaptic maturation as well.

Resting neural activity patterns in auditory brainstem and midbrain in conductive hearing loss

Conclusions: Conductive hearing loss (CHL) lowers resting neural activity patterns in the auditory periphery. Such reductions of peripheral auditory activity may influence the developing central brain during early postnatal years when the system is still highly plastic. Objectives: A common cause of CHL in young children is otitis media; if chronic and/or episodic there may be a risk to speech and language development. In this clinical context we have investigated changes in neural activity patterns in the brainstem and midbrain in an animal model of CHL. Methods: In a mouse model, a 50–60 dB CHL was produced by blocking the ear canals. We quantified resting neural activity patterns in the cochlear nucleus and inferior colliculus using c-fos immuno-labelling. This experimental group was compared with normal-hearing controls and with animals with bilateral cochlear ablation. Results: Subjects with CHL had a statistically significant reduction in c-fos-labelled cells in the cochlear nucleus and central nucleus of the inferior colliculus compared with normal controls. This decreased c-fos expression suggests a change in resting neural activity generated at the inner hair cell synapse, leading to a reduction in activity levels in the ascending auditory pathways.

Exocytosis in the Frog Amphibian Papilla

Using whole-cell patch-clamp recordings, we measured changes in membrane capacitance (ΔC (m)) in two subsets of hair cells from the leopard frog amphibian papilla (AP): the low-frequency (100-500 Hz), rostral hair cells and the high-frequency (500-1200 Hz), caudal hair cells, in order to investigate tonotopic differences in exocytosis. Depolarizations of both rostral and caudal hair cells evoked robust ΔC (m) responses of similar amplitude. However, the calcium dependence of release, i.e., the relationship between ΔC (m) relative to the amount of calcium influx (Q (Ca) (2+)), was found to be linear in rostral hair cells but supra-linear in caudal hair cells. In addition, the higher numbers of vesicles released at caudal hair cell active zones suggests increased temporal precision of caudal hair cell exocytosis. ΔC (m) responses were also obtained in response to sinusoidal stimuli of varying frequency, but neither rostral nor caudal hair cell ΔC (m) revealed any frequency selectivity. While all AP hair cells express both otoferlin and synaptotagmin IV (SytIV), we obtained evidence of a tonotopic distribution of the calcium buffer calretinin which may further increase temporal resolution at the level of the hair cell synapse. Our findings suggest that the low (rostral) and high (caudal) frequency hair cells apply different mechanisms for fine-tuning exocytosis.

Sharp Ca2+Nanodomains beneath the Ribbon Promote Highly Synchronous Multivesicular Release at Hair Cell Synapses

Hair cell ribbon synapses exhibit several distinguishing features. Structurally, a dense body, or ribbon, is anchored to the presynaptic membrane and tethers synaptic vesicles; functionally, neurotransmitter release is dominated by large EPSC events produced by seemingly synchronous multivesicular release. However, the specific role of the synaptic ribbon in promoting this form of release remains elusive. Using complete ultrastructural reconstructions and capacitance measurements of bullfrog amphibian papilla hair cells dialyzed with high concentrations of a slow Ca²⁺ buffer (10 mM EGTA), we found that the number of synaptic vesicles at the base of the ribbon correlated closely to those vesicles that released most rapidly and efficiently, while the rest of the ribbon-tethered vesicles correlated to a second, slower pool of vesicles. Combined with the persistence of multivesicular release in extreme Ca²⁺ buffering conditions (10 mM BAPTA), our data argue against the Ca²⁺-dependent compound fusion of ribbon-tethered vesicles at hair cell synapses. Moreover, during hair cell depolarization, our results suggest that elevated Ca²⁺ levels enhance vesicle pool replenishment rates. Finally, using Ca²⁺ diffusion simulations, we propose that the ribbon and its vesicles define a small cytoplasmic volume where Ca²⁺ buffer is saturated, despite 10 mM BAPTA conditions. This local buffer saturation permits fast and large Ca²⁺ rises near release sites beneath the synaptic ribbon that can trigger multiquantal EPSCs. We conclude that, by restricting the available presynaptic volume, the ribbon may be creating conditions for the synchronous release of a small cohort of docked vesicles.

Both pre- and postsynaptic activity of Nsf prevents degeneration of hair-cell synapses.

Vesicle fusion contributes to the maintenance of synapses in the nervous system by mediating synaptic transmission, release of neurotrophic factors, and trafficking of membrane receptors. N-ethylmaleimide-sensitive factor (NSF) is indispensible for dissociation of the SNARE-complex following vesicle fusion. Although NSF function has been characterized extensively in vitro, the in vivo role of NSF in vertebrate synaptogenesis is relatively unexplored. Zebrafish possess two nsf genes, nsf and nsfb. Here, we examine the function of either Nsf or Nsfb in the pre- and postsynaptic cells of the zebrafish lateral line organ and demonstrate that Nsf, but not Nsfb, is required for maintenance of afferent synapses in hair cells. In addition to peripheral defects in nsf mutants, neurodegeneration of glutamatergic synapses in the central nervous system also occurs in the absence of Nsf function. Expression of an nsf transgene in a null background indicates that stabilization of synapses requires Nsf function in both hair cells and afferent neurons. To identify potential targets of Nsf-mediated fusion, we examined the expression of genes implicated in stabilizing synapses and found that transcripts for multiple genes including brain-derived neurotrophic factor (bdnf) were significantly reduced in nsf mutants. With regard to trafficking of BDNF, we observed a striking accumulation of BDNF in the neurites of nsf mutant afferent neurons. In addition, injection of recombinant BDNF protein partially rescued the degeneration of afferent synapses in nsf mutants. These results establish a role for Nsf in the maintenance of synaptic contacts between hair cells and afferent neurons, mediated in part via the secretion of trophic signaling factors.

Harmonin inhibits presynaptic Cav1.3 Ca2+ channels in mouse inner hair cells

Harmonin is a scaffolding protein required for normal mechanosensory function in hair cells. Here, we describe a novel presynaptic association of harmonin and Cav1.3 Ca2+ channels at the mouse inner hair cell synapse, which limits channel availability through a ubiquitin-dependent pathway.

What the cochlea does next : encoding sound at the cochlear inner hair cell synapse

What the cochlea does next : encoding sound at the cochlear inner hair cell synapse

Recovery from Short-Term Depression and Facilitation Is Ultrafast and Ca2+Dependent at Auditory Hair Cell Synapses

Short-term facilitation and depression coexist at many CNS synapses. Facilitation, however, has not been fully characterized at hair cell synapses. Using paired recordings and membrane capacitance measurements we find that paired-pulse plasticity at an adult frog auditory hair cell synapse depends on pulse duration and interpulse intervals. For short 20 ms depolarizing pulses, and interpulse intervals between 15 and 50 ms, facilitation occurred when hair cells were held at -90 mV. However, hair cells held at -60 mV displayed only paired-pulse depression. Facilitation was dependent on residual free Ca2+ levels because it was greatly reduced by the Ca2+ buffers EGTA and BAPTA. Furthermore, low external Ca2+ augmented facilitation, whereas depression was augmented by high external Ca2+, consistent with depletion of a small pool of fast releasing synaptic vesicles. Recovery from depression had a double-exponential time course with a fast component that may reflect the rapid replenishment of a depleted vesicle pool. We suggest that hair cells held at more depolarized in vivo-like resting membrane potentials have a tonic influx of Ca2+; they are thus in a dynamic state of continuous vesicle release, pool depletion and replenishment. Further Ca2+ influx during paired-pulse stimuli then leads to depression. However, at membrane potentials of -90 mV, ongoing release and pool depletion are minimized, so facilitation is revealed at time intervals when rapid vesicle pool replenishment occurs. Finally, we propose that vesicle pool replenishment kinetics is not rate limited by vesicle endocytosis, which is too slow to influence the rapid pool replenishment process.

Molecular Anatomy of the Hair Cell's Ribbon Synapse

Hearing depends on reliable and temporally precise neurotransmission by cochlear hair cells. The wide dynamic range and high sensitivity with which these cells encode acoustic stimuli are associated with a presynaptic specialization termed the presynaptic dense body or synaptic ribbon. Apposed to the presynaptic density, this spherical or flattened structure tethers a layer of synaptic vesicles and is thought to facilitate their exocytotic fusion. Although defining the molecular constituents of the hair cell's synaptic ribbon should contribute to our understanding of neurotransmitter release at this synapse, accomplishing this task has been slowed by the difficulty of obtaining sufficient amounts of starting material for protein analysis from hair cells. We isolated synaptic material from chicken cochleas, purified synaptic ribbons with specific immunological reagents, and identified the associated proteins by tandem mass spectrometry. Purification of the ribbons revealed a predominant composition of C-terminal-binding proteins, especially ribeye, in association with the small GTPase Rab3, which is possibly involved in attaching vesicles to the ribbon. In comparison with the components of conventional synapses and of retinal ribbon synapses, we observed that certain regulatory proteins are excluded from the hair cell's synapse. Using antisera against several of the novel proteins and membrane-trafficking components that we had identified, we documented their localization in isolated hair cells. Our results indicate that the ribbon synapses of hair cells display modifications to the presynaptic machinery that are associated with the high-fidelity transmission of acoustic signals to the brain.

Ca2+and Ca2+-Activated K+Channels That Support and Modulate Transmitter Release at the Olivocochlear Efferent–Inner Hair Cell Synapse

In the mammalian auditory system, the synapse between efferent olivocochlear (OC) neurons and sensory cochlear hair cells is cholinergic, fast, and inhibitory. This efferent synapse is mediated by the nicotinic alpha9alpha10 receptor coupled to the activation of SK2 Ca(2+)-activated K(+) channels that hyperpolarize the cell. So far, the ion channels that support and/or modulate neurotransmitter release from the OC terminals remain unknown. To identify these channels, we used an isolated mouse cochlear preparation and monitored transmitter release from the efferent synaptic terminals in inner hair cells (IHCs) voltage clamped in the whole-cell recording configuration. Acetylcholine (ACh) release was evoked by electrically stimulating the efferent fibers that make axosomatic contacts with IHCs before the onset of hearing. Using the specific antagonists for P/Q- and N-type voltage-gated calcium channels (VGCCs), omega-agatoxin IVA and omega-conotoxin GVIA, respectively, we show that Ca(2+) entering through both types of VGCCs support the release process at this synapse. Interestingly, we found that Ca(2+) entering through the dihydropiridine-sensitive L-type VGCCs exerts a negative control on transmitter release. Moreover, using immunostaining techniques combined with electrophysiology and pharmacology, we show that BK Ca(2+)-activated K(+) channels are transiently expressed at the OC efferent terminals contacting IHCs and that their activity modulates the release process at this synapse. The effects of dihydropiridines combined with iberiotoxin, a specific BK channel antagonist, strongly suggest that L-type VGCCs negatively regulate the release of ACh by fueling BK channels that are known to curtail the duration of the terminal action potential in several types of neurons.