Elsevier

Hearing Research

Volume 373, 1 March 2019, Pages 85-95
Hearing Research

Research Paper
Ontogeny of auditory brainstem responses in the bat, Phyllostomus discolor

https://doi.org/10.1016/j.heares.2018.12.010Get rights and content

Highlights

  • We present auditory brainstem responses, a minimally invasive technique to assess basic hearing in echolocating bats.

  • We show that P. discolor bats have good hearing well into the ultrasonic range already very early in life, 7 days after birth.

  • Hearing improves further until about one month of age.

  • Thereupon the audiogram becomes more structured, reflecting the growth of outer ear and tragus.

  • ABRs decrease in amplitude and shorten with age, correlated with animal growth and refinement of fast auditory processing.

Abstract

Hearing is the primary sensory modality in bats, but its development is poorly studied. For newborns, hearing appears essential in maintaining contact with their mothers and to develop echolocation abilities. Here we measured auditory brainstem responses (ABRs) to clicks and narrowband tone pips covering a large frequency range (5–90 kHz) in juveniles (p7 to p200) and adults of the bat, Phyllostomus discolor. Tone-pip audiograms show that juveniles at p7 are already quite responsive, not only below 20 kHz but up to 90 kHz. Hearing sensitivity increases further until about p14 and is then refined, possibly correlated with growth and differentiation of the animals’ outer ears. ABR amplitudes decrease within the first 3–4 months, inversely correlated with the bat weight and forearm length. ABR Wave I latency decreases with increasing stimulation level. ABR duration (measured between Waves I and V) is longer in juveniles and shortens with age which may reflect temporal refinement of auditory brainstem neurons to accommodate the exceptional temporal precision required for effective echolocation. Overall our data show that P. discolor bats have good hearing very early in life. The current method represents a fast and minimally invasive way of characterizing basic hearing in bats.

Introduction

Bats are nocturnal animals with little natural illumination in their environment. Consequently, hearing is their primary sensory modality. Various acoustic signals are used for communication. But bats also orient with acoustic signals: bats echolocate by emitting ultrasonic signals and listen to reflections of obstacles in their environment.

Hearing can be studied, among other methods, by recording non-invasively the auditory brainstem response (ABR) to acoustic stimuli via electrodes placed on the scalp of an anesthetized animal. The ABR reflects the summed electrical responses of both the auditory nerve fibers and the nuclei of the central auditory pathway (Burkard et al., 2007). The strongly transient stimulation for ABRs bears similarities to the transient acoustic stimulation by calls emitted by bats when engaged in echolocation. The ABR waveform contains several identifiable oscillations, labeled usually with roman numerals. It is commonly accepted that wave I represents the compound response from the auditory nerve while the later waves represent responses from the ascending auditory pathway. ABRs are a fast and minimally invasive method to test hearing of a study subject. One has to keep in mind though, when measuring auditory thresholds by means of ABRs, these ABR thresholds refer only to the lowest sound pressure level (SPL) that can generate identifiable electrical response waves and not to the actual hearing threshold. For animals, behaviorally obtained hearing thresholds are often several tens of dB lower than the ABR thresholds (Yuen et al., 2005).

Recordings of ABRs are standard methods for principal hearing measurements in humans (Burkard et al., 2007) and many animals (Corwin et al., 1982). However, in bats this has only been applied a handful of times during the last 60 years e.g. Myotis lucifugus (Grinnell, 1963), Plecotus townsendii (Grinnell, 1963), Rousettus aegyptiacus (Belknap et al., 1982), Noctilio leporinus (Wenstrup, 1984), Lasiurus borealis (Obrist et al., 1998), Eptesicus fuscus (Burkard et al., 1994; Simmons et al., 1990), and Pipistrellus abramus (Boku et al., 2015; Simmons et al., 2015).

The shape of the ABR waveform changes progressively with age and reflects the maturation of auditory processing of the animal. In juveniles these changes reflect the maturation of neural pathways from the inner ear through the brainstem towards the auditory cortex. In older individuals these changes can reflect degradation or lesions of the auditory pathway. Such changes are quantified in parameters like wave amplitudes, wave latencies and inter-wave latencies. Therefore, ABRs are an effective method to study basic principles of the ontogeny of the auditory pathway. Further, it allows observing the development within the same individual over extended developmental periods. While this can also be done with e.g. otoacoustic emissions, ABRs allow some conclusions also of auditory maturation beyond the inner ear. In classical electrophysiological studies where hearing is characterized from extracellular neural responses (Brown et al., 1978, 1980; Sterbing, 2002), different individuals at different ages are looked at because most neurological investigations are invasive and animals are euthanatized for accompanying histological preparations at the end. Behavioral auditory studies, on the other hand, are difficult to implement in juveniles because it takes usually an extended period of time to train a testing paradigm during which no data collection is possible. Thus, behavioral data on hearing cannot be obtained from very young animals.

Many model animals in hearing research are born deaf and undergo rapid maturation of the auditory periphery throughout the 2nd week of life. In contrast e.g. humans have a well-developed auditory system at birth and even pre-birth (Berkson et al., 1974; Birnholz et al., 1983). In bats both developmental models exist. Bat species like Myotis oxygnathus (Konstantinov, 1973), Rhinolophus ferrrumequinum (Konstantinov, 1973), Rhinolophus rouxi (Rubsamen, 1987) Hipposideros speoris (Rubsamen et al., 1989), Myotis velifer (Brown et al., 1980), and Antrozous pallidus (Brown et al., 1978) were described to be deaf at birth and gradually develop their hearing during the first weeks of life. Hearing onset of e.g. Antrozous pallidus is around p7 and these juveniles are only sensitive to frequencies between 5 and 15 kHz. During the following weeks the upper frequency limit gradually increases up to about 90–100 kHz and the auditory system becomes more sensitive in general (Brown et al., 1978). Similar to humans, other bat species like Peteronotus parnellii (Brown et al., 1980), Pteronotus suapurensis (Brown et al., 1980; Vater et al., 2003) and Carollia perspicillata (Sterbing, 2002) have good hearing already at birth. Carollia perspicillata is already sensitive from birth at frequencies between 15 and 30 kHz and after one week up to 77 kHz. Within seven weeks of development, the bats sensitive frequency range expands up to 110 kHz and becomes gradually more sensitive at all frequencies (Sterbing, 2002). The target species of the current study, Phyllostomus discolor, has been shown to react to maternal directive calls within 24 h of birth (Esser et al., 1990). However, other than that, research on hearing in this species has focused on adults. P. discolor hearing covers a frequency range up to at least 100 kHz (Hoffmann et al., 2008a). However, neural and behavioral audiograms exist only of adult individuals; the development of the auditory system in juveniles has not been investigated.

Here, we report on the hearing development of P. discolor juveniles between p7 and p200 and compare the results to adults. Data were collected by means of ABR and allowed us to repeatedly test the same individuals at different ages. We report details of the ABR waveform changes and the overall hearing development. This data arose as part of a larger project on vocal development in normal hearing and deafened juveniles.

Section snippets

Animals and experimental approval

All animals are part of a breeding colony housed at the Department of Neurobiology at Ludwig-Maximilians University, Munich, Germany. Hearing tests were performed with six juveniles for the first time at ages between p7 and p11. Thereafter three of the juveniles were tested every week for the first months, every second week for month two and three, and once a month for additional three months. As a small control group, two extra juveniles were tested only once at the age of p200. Eight adults

Results

We measured significant ABRs from six juveniles with a maximum age of p11, repeatedly of three juveniles (up to 12 times) within the first six months of their life, of two sub-adults at the age of p200, and of eight adults of unknown age ≥ 2 years. We obtained ABR audiograms using tone pip stimuli and click evoked ABRs to broadband impulses for all individuals.

Discussion

The auditory brainstem response is well-known as an effective and minimally invasive tool to assess basic auditory function (Burkard et al., 2007). Therefore ABRs allow monitoring the development of auditory function over time in the same individual (Brandt et al., 2013; Kraemer et al., 2017). As opposed to behavioral audiograms, which in animal models often require weeks or months of training and data acquisition, an ABR threshold can be measured from an anesthetized animal within about 1 min

Author contributions

ML and LW conceived the study.

ML conducted the study and analyzed the data.

ML and LW wrote the manuscript.

Conflicts of interest

The authors declare that they have no conflict of interest.

Acknowledgments

We are thankful for the help from Uwe Firzlaff in initially establishing ABR recordings, from Vanessa Kratzer and Ella Lattenkamp during ABR recordings and the valuable discussions on the data and manuscript with the members of the Division of Neurobiology lead by Benedikt Grothe. This work was funded by a Human Frontier Science Program (HFSP) Research Grant (RGP0058/2016) awarded to Lutz Wiegrebe and colleagues.

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