Elsevier

Brain Research

Volume 1148, 7 May 2007, Pages 83-89
Brain Research

Research Report
Creatine and tempol attenuate noise-induced hearing loss

https://doi.org/10.1016/j.brainres.2007.02.021Get rights and content

Abstract

To define the role of free radical formation and potential energy depletion in noise induced hearing loss (NIHL), we measured the effectiveness of tempol (free radical scavenger) and creatine (enhances cellular energy storage) alone and in combination to attenuate NIHL. Guinea pigs were divided into four treatment groups: controls, 3% creatine diet (2 weeks prior to noise exposure), tempol (3 mM in drinking water 2 weeks prior to exposure), and creatine plus tempol and exposed to 120 dB SPL one-octave band noise centered at 4 kHz for 5 h. The noise-only control group showed frequency-dependent auditory threshold shifts (measured by auditory brainstem response, ABR) of up to 73 dB (16 kHz) on day 1, and up to 50 dB (8 kHz) on day 10. Creatine-treated subjects had significantly smaller ABR threshold shifts on day 1 and on day 10. Tempol alone significantly reduced ABR threshold shifts on day 10 but not on day 1. ABR shifts after combination treatment were similar to those in the creatine group. Hair cell loss on day 10 was equally attenuated by creatine and tempol alone or in combination. Our results indicate that the maintenance of ATP levels is important in attenuating both temporary and permanent NIHL, while the scavenging of free radicals provides protection from permanent NIHL.

Introduction

Exposure to high-intensity sound triggers a decrease in local blood flow in the inner ear, especially at very high sound exposure levels of 120–155 dB (Perlman and Kimura, 1962, Quirk et al., 1992, Miller et al., 1996), while metabolic activity remains elevated well above basal levels (Canlon and Schacht, 1983). At very high sound exposure levels, the reduction in cochlear blood flow (CBF) and local vasoconstriction can subject the cochlea to severe hypoperfusion in the presence of high energy demands. Impaired energy status can lead to activation of excitatory amino acid receptors (Brown and Borutaite, 2002), increasing intracellular calcium (White et al., 2000), and the generation of free radicals (Lang-Rollin et al., 2003), which are all potentially damaging to the inner ear. If the initial step in the development of noise trauma is a depletion of cellular energy stores, then agents that can buffer cellular energy stores may be protective. Creatine kinase is a key enzyme involved in regulating energy metabolism in cells with intermittently high and fluctuating energy requirements (Weiss et al., 2005), including the inner ear (Spicer and Schulte, 1992). The enzyme catalyzes the transfer of high-energy phosphate from phosphocreatine to ADP to generate ATP. Several cytoplasmic and mitochondrial isoforms have been identified that, with the substrates creatine and phosphocreatine, constitute an intricate cellular energy buffering and transport system connecting sites of energy production to sites of energy consumption (Hemmer and Wallimann, 1993). We tested the hypothesis that compounds that increase the cochlear energy reserve will be protective by assessing if oral administration of creatine attenuates noise-induced hearing loss.

Another putative cause of noise-induced hair cell death is oxidative stress. The formation of reactive oxygen species (ROS) results from high levels of mitochondrial activity induced by noise (Balaban et al., 2005, Gourlay and Ayscough, 2005, Yamashita et al., 2004, Ohinata et al., 2000), and is exacerbated by reduction in CBF, which accompanies intense sound exposure. Consequently, ROS scavengers and inhibitors attenuate NIHL (Yamasoba et al., 1999, Henderson et al., 1999, Kopke et al., 2000, Ohinata et al., 2003). Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) is a water-soluble analogue of the spin label TEMPO, widely employed in electron spin resonance spectroscopy (Saito et al., 2003). Tempol permeates biological membranes and acts as a spin trap for superoxide radicals. It can reduce superoxide-related injury in ischemia/reperfusion, inflammation, and radiation-induced damage (Thiemermann, 2003). In the present study we assessed the efficacy of tempol to prevent NIHL.

Creatine and tempol should act on different pathways associated with NIHL. Several studies have shown that an additive protection against NIHL can be achieved by combining several agents acting against different damaging mechanisms (Hight et al., 2003, Kopke et al., 2000, Yamashita et al., 2005; for recent reviews, see Miller et al., 2006 and Le Prell et al., 2006). To provide further insights into the independence of these factors and the potential efficacy of a combination of agents, we examine the effectiveness of creatine or tempol alone and in combination on auditory threshold shifts and hair cell damage induced by noise exposure.

Section snippets

ABR

Temporary noise-induced threshold shifts (TTS), measured one-day post-noise exposure, were significantly reduced by treatment with creatine, or a combination of creatine plus tempol. Permanent noise-induced hearing loss measured 10 days post-noise was significantly reduced by treatment with creatine, tempol, or a combination thereof at 4 and 8 kHz (see Fig. 1). Only the combination of creatine and tempol provided a significant attenuation of the noise-induced threshold shift at 4 kHz 10 days

Discussion

Creatine can reduce both temporary (TTS) and permanent (PTS) threshold shifts, while the antioxidant tempol attenuates the PTS only. At 16 kHz we also observed that the combination of these two agents provided a significantly greater attenuation of the noise-induced threshold shift than tempol alone. This additivity, however, was not observed at lower test frequencies and was not supported by histological assessment.

The creatine-mediated reduction in both temporary and permanent ABR threshold

Experimental design

Pigmented male guinea pigs (200–400 g, 2–4 weeks old; Elm Hill Breeding Labs, Chelmsford, MA) were used in this study. Creatine and tempol were obtained from Sigma (St. Louis, MO). Creatine was administered orally to the guinea pigs in their food at doses of 3% of the diet. Tempol was administered at 3 mM in drinking water. Controls received unsupplemented but otherwise identical diets and drinking water. The diets were administered over a total of 24 days beginning 14 days before noise

Acknowledgments

Supported by NIH DC 04058 and DC 05188, General Motors Corp., United Autoworkers Union, and the Ruth and Lynn Townsend Professorship of Communication Disorders. We wish to thank Hollie Valentine for her technical assistance.

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