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Auditory Cortical Cells with Electrode in Place
Auditory Neuroscience Laboratory
Dept. of ASC
College of COMM Arts & Sciences

Effects of Lead on Biological Systems

Introduction
We have an interest on the effects of lead on children, particularly as it relates to the auditory system, and whether lead causes hearing loss or deafness. Our research studies are conducted at the organismic level and at the cellular level. At the organismic level, we utilized auditory assessment methods such as Video Otoscopy, Real Ear (acoustic characteristics of the ear canal), Middle Ear Impedance, Otoacoustic Emissions, Auditory Brain Stem Evoked Responses, and Late Auditory Evoked Potentials. For our cellular studies, we utilized combined techniques of Cell Culture, Patch Clamp Electrophysiology, Immunocytochemistry, Photomicrography, and Molecular Biology.
Description and Background of Lead
The normal development of the young central nervous system (CNS) is vulnerable to a number of environmental factors, including lead (Pb2+). Harmful levels of lead have concerned educators and researchers for a number of years due to its debilitating effect on learning and memory. Pb2+ exists in the air, drinking water, soil, wall fixtures, dust, lead-based paints, venetian blinds and several industrial products. It is a potent environmental toxicant that has been associated with adverse CNS and behavioral disorders of both laboratory animals and humans. Surveys conducted by the Public Health Service concluded that environmental Pb2+ poisoning is among the top four diseases of young children. Pb2+ is so potent that the Center for Disease Control has had to lower its risk for neurotoxicity several times, from blood-lead levels of 60 mg/dl to the current accepted level of 10 mg/dl.

The majority of research that has examined the effects of Pb2+ exposure has identified children exhibiting deficits of the CNS. Children exposed to high levels of Pb2+ prenatally and others who were exposed to Pb2+ postnatally exhibited poorer scores on measures of cognitive function and central auditory processing abilities. Pb2+ -induced impairments are of concern because they are believed to lead to learning and memory disorders. Some researchers report that Pb2+ intoxication is most intense during the first two years of life, suggesting a critical period for impairment. The period coincides with the critical period for normal language and speech development.

Some investigators previously believed that Pb2+ exposure posed more of a threat to the young developing CNS than to the developed CNS system. However, it has been reported that the cochlear nerve and central auditory structures were preferentially sensitive to the effects of Pb2+ exposure in both developing and mature systems. In fact, elevated hearing thresholds in both young children and adults and increased latencies of auditory brainstem responses in adults have been reported as a result of exposure to environmental Pb. Cases of occupationally exposed workers with acute and chronic episodes of Pb2+ intoxication have been investigated. The highest levels of blood-Pb2+ levels ranged from 40 mg/dl to 175 mg/dl, with a mean of 83 mg/dl. Abdominal pain, headache and fatigue were among the many reported symptoms among the population.

Limited information is available concerning the effects of Pb2+ on sensory function and the mechanism(s) responsible for lead toxicity. However, sufficient data are available which support the occurrence of elevated hearing threshold levels, as well as an increase in the threshold of activation for the initiation of an action potential in the presence of varying levels of Pb2+. In addition, histological studies of auditory structures have found segmental demyelinization and axonal degeneration of the cochlea nerve with no apparent effect on the vestibular and spiral ganglia. Unlike other voltage-gated ionic channels (e.g., Ca2+) and ligand-gated ionic channels, the question still remains, do hair cells or spiral ganglion cells (SGCs) of the inner ear exhibit normal pharmacological and kinetic channel activity in the presence of Pb2+ within the mammalian auditory system?

In an attempt to identify the mechanism(s) responsible for Pb2+ neurotoxicity, pharmacological studies have investigated the effects of Pb2+ on synaptic transmitter release, kinetic properties of ionic currents and sensitivity to neuronal development. A group of investigators in 1984 studied neurotransmitter release at the neuromuscular junction of rats and found that Pb2+ (100 mM) effectively blocked the nerve-evoked end-plate potential (EPP). Both effects were reversible when washed with normal external solutions. Another investigator in 1990 investigated the effects of Pb2+ on two receptor operated ion channels [Serotonin or 5-HT and neuronal nicotinic acetylcholine (ACh) receptors] and two voltage-gated ion channels (Na+ and Ca2+) of mouse neuroblastoma cells. The ACh induced inward current was blocked by nanomolar concentrations of Pb2+. Comparatively, the transient inward current mediated by 5-HT receptors required a larger quantity of Pb2+ (1.0 mM) before a steady reduction of current amplitude was observed. High concentrations of barium (Ba2+) produced a fast transient as well as a noninactivating inward current. Both types of currents were reduced in the presence of 10 mM of Pb2+. However, the Na+ current remained unaffected up to 100 mM of Pb2+.

Abnormalities in the movement, distribution, as well as inhibition in the functional capacity of Ca2+ channels in the CNS due to Pb2+ neurotoxicity may play an important role in neuronal development. One investigator found that chronic exposure of rats to low levels of Pb2+ reduced the number of inositol triphosphate (IP3) second messenger receptors and thus reduced the capacity of IP3 to mobilize Ca2+ from intracellular sources. This action may alter normal cell development and thus lead to CNS dysfunction.

The NMDA glutamatergic receptor is known to be involved in the processes of learning and plasticity as well as playing a key role in the early stages of neuronal development. Thus, it was demonstrated that Pb2+ blocked NMDA induced currents of cultured hippocampal neurons in an age-dependent manner. In other words, 10 mM Pb2+ decreased the NMDA response by 57% during the first week; however, the same concentration of Pb2+ during the fourth week of exposure only reduced the NMDA response by 15%.

Two other investigators examined the response of divalent cations (Ca2+, Pb2+, Cd2+) on both tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium channels of rat dorsal root ganglion (DRG) cells. The peak TTX-S current was reduced by 20% in the presence of externally applied Pb2+ compared to 22% by the addition of 5 m M Ca2+. The activation voltage was unaffected by Ca2+ but shifted in the depolarized direction in the presence of Pb2+ (50 mM). Conduction-voltage curves showed a minimal decrease in the maximal conductance of the TTX-S currents in the presence of Pb2+. Peak TTX-R currents were more sensitive to the actions of Pb2+ and Cd2+ and less sensitive to Ca2+ than TTX-S currents. The TTX-R current was reduced by 70.4% at a membrane level of +10mV by 50 mM Pb2+. Conductance curves were measured at membrane potentials where conductance attained 50% of its maximal values. Conductance curves were shifted 25.3 mV by Pb2+. Maximal conductance decreased 55.2% to Pb2+. Effects seen for both TTX-S and TTX-R were reversible after washing the cell for 1 - 2 minutes with normal external solution.

Pharmacology of Lead
Pb2+ is one of several heavy metals that is toxic to the body and found to cause lead poisoning. It exerts its toxic effect by combining with one or more reactive groups (ligands) essential for normal physiological functions. Heavy metals such as Pb2+ react in the body with ligands containing oxygen and nitrogen. This creates a metal complex formed by a coordinate bond. Heavy metal antagonists known as chelating agents are designed to reverse the binding of Pb2+ to body ligands by blocking the binding of Pb2+ to the ligands. One investigator defines a chelate as a complex formed between a metal and compound that contains two or more potential ligands. This reaction produces a heterocyclic ring. Chelating agents are designed to compete with the groups of ligands for the metal, thereby, preventing or reversing the toxic effects and enhancing the excretion of the metals.

Pb2+ poisoning can be either acute or chronic. Acute Pb2+ poisoning occurs infrequently and typically results from ingestion of acid-soluble Pb2+ compounds or inhalation of Pb2+ vapors. Chronic Pb2+ poisoning, also known as plumbism, can have adverse affects involving one or any combination of the following systems: gastrointestinal system, neuromuscular system, CNS, hematological and renal systems. The absorption of Pb2+ into the body enters by two major routes, the respiratory and the gastrointestinal tract. Absorption of inhaled Pb2+ depends upon the form of Pb2+ (vapor vs. particles) ingested as well as the concentration of ingested Pb2+. Ninety nine percent of Pb2+ absorbed into the blood stream binds to hemoglobin in erythrocytes. Inorganic Pb2+ is initially distributed in soft tissue organs such as the liver and kidneys. It is later found redistributed and deposited in bone, teeth (seen as lead lines) and hair. Only small amounts of inorganic Pb2+ accumulate in the white matter of the brain, with most of the deposited Pb2+ found in gray matter and the basal ganglia.

N.B.: Several research papers and key references were consulted in order to write this review. We have chosen not to list the references here for the sake of brevity. A version of this review is available with references and bibliography. To obtain a copy, please contact mooreer@pilot.msu.edu and it will be downloaded to your computer within a reasonable time.

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1998 Dr. Ernest J. Moore, Ph.D.
all rights reserved