Introduction

 Amyotrophic Lateral Sclerosis

Amyotrophic Lateral Sclerosis (ALS,) also known as Lou Gehrig’s disease, is a degenerative neurological disease characterized by muscle atrophy and paralysis.  This motor neuron disease is progressive and chronic, wherein the spinal cord neurons responsible for supplying electrical stimulation to the muscles die. As a result of motor neuron death, muscles fail to receive stimulation and therefore deteriorate.  There are two major questions relating to research of ALS; the first being the cause of motor neuron death, and the second being possible treatment of the disease.  One possible treatment option is the replacement of motor neurons with differentiated stem cells.  Through specific treatment, embryonic stem cells can be forced to differentiate along a motor neuron line taking on structure characteristic of motor neurons. In determining whether these differentiated stem cells have the capacity to act as motor neurons, three factors must be addressed: (1) can the cells receive impulses; (2) can the cells appropriately interpret their inputs and transform these to appropriate outputs; and (3) can the cells send impulses to the appropriate targets?  One key to the ability of the second factor above is the presence of voltage-gated calcium channels.

 The Nervous System

The nervous system is the most complicated and highly organized system of the human body. The nervous system is responsible for receiving, sorting, and transmitting nerve signals throughout the body and can be divided into two parts, central and peripheral.  The central nervous system consists of the brain and the spinal cord, while the peripheral nervous system consists of a series of nerves by which the central nervous system is connected with the various tissues of the body.  The spinal cord contains complex networks of neurons, some of which are responsible for the production of movement. The “final common pathway” of output from the spinal cord is the motor neuron. The cell bodies of motor neurons reside in the ventral horn of the spinal cord, and their axons leave via the ventral root to innervate all of the skeletal muscles of the body.

 The Neuron

The neuron is the basic unit of the nervous system; there are a number of different classifications of neurons, including sensory neurons, inter neurons and motor neurons.  Motor neurons are those neurons whose role is specifically to stimulate muscular action.  All nerve cells consist of dendrites leading to a cell body.  This cell body is connected to a long axon insulated with myelin terminating in axon terminals, which release neurotransmitters across the synapse.  Nerve cells receive and transmit impulses through a series of electrical ‘transactions.’  A resting neuron is electrically charged because of the uneven distribution of sodium, potassium and chloride ions across the cell membrane.  The interior of the cell is more electronegative than the exterior, resulting in the cell having a charge of approximately –70 millivolts, caused by the selective permeability of the membrane to potassium ions.  The stimulation of a neuron either mechanically, or chemically (by neurotransmitters) results in the depolarization of the cell.  The resting potential is decreased to about –50 millivolts by the transfer of sodium ions across the plasma membrane into the cell.  Once this threshold potential is reached, voltage-gated sodium channels allow thousands of sodium ions to rush into the cell, which in turn open voltage-gated sodium channels further down the axon.  This is the action potential, also known as the transmission of the nerve impulse. In the cell soma and dendrites, the depolarization of the action potential leads to the opening of many different classes of calcium channel, which in turn leads to calcium influx. Amongst other effects, this leads to an enhancement of the depolarization. This enhancement is critical to normal motor neuron function and locomotion.    

 Voltage-gated Ion Channels

Ion channels are present in the membranes that surround all cells.  By conducting and controlling the flow of ions, these pore-forming enzymes help establish the small negative voltage that all cells possess at rest.  An ion channel is an integral membrane protein or group of proteins which either span or are sufficiently embedded in the membrane with which they are associated.  Such "multi-subunit" assemblies usually involve a circular arrangement of identical or related proteins closely packed around a water-filled pore through the plane of the membrane or lipid bilayer.  This type of channel pore is just one or two atoms wide at its narrowest point, it conducts a specific species of ion, such as calcium.   Access to the pore is governed by "gates," which may be opened or closed by chemical or electrical signals, or mechanical force, depending on the variety of channel.

 Voltage-gated channels, or those whose access is governed by electrical signals, underlie the nerve impulse.  Ion channels figure in a wide variety of biological processes that involve rapid changes in cells. Voltage-gated channels sense the transmembrane potential and open or close in response to depolarization or hyperpolarization, respectively. Examples include the sodium and potassium voltage-gated channels of nerve and muscle, and the voltage-gated calcium channels that control neurotransmitter release in presynaptic endings.

 The voltage-gated channels that underlie the nerve impulse consist of four subunits with six transmembrane helices each.  Upon activation, these helices move about and open the pore. Two of these six helices are separated by a loop that lines the pore and is the primary determinant of ion selectivity and conductance in this channel class and some others.  The L-type voltage-gated calcium channels is a cell membrane glycoprotein forming a channel in a biological membrane selectively permeable to calcium ions. Calcium is essential for a variety of bodily functions, such as neurotransmission, muscle contraction and proper heart function.  These voltage dependent calcium channels have multiple subunits; alpha-1, alpha-2, beta and delta in a 1:1:1:1 ratio.  Calcium channels are the main cause of action potentials in certain smooth muscles, assist in hyperpolarization and in stimulating intracellular processes, such as the release of neurotransmitters.

Stem Cells

A stem cell is a cell that is described as multipotent; that is, one that gives rise to a lineage of cells.  Upon division, the mother cell produces dissimilar daughters, one replacing the original stem cell, the other differentiating further.  Inductive signals and transcription factors involved in motor neuron (MN) generation have been identified, and it has been shown that developmentally relevant signaling factors can induce mouse embryonic stem (ES) cells to differentiate into spinal progenitor cells and motor neurons very similar to in vivo development.  Inductive signals involved in normal pathways of neurogenesis can direct ES cells to form specific classes of Central Nervous System neurons; these cells can populate embryonic spinal cords, extend axons and form synapses with target muscles. 

 Spinal motor neurons are one central nervous system neuronal subtype for which pathways of neuronal specification have been defined.  First, ectodermal cells acquire an initial rostral neural character through the regulation of BMP, FGF and Wnt signaling.  They acquire spinal positional identify in response to caudalizing signals that include retinoic acid (RA.)  The spinal progenitor cells acquire a MN progenitor identity in response to the ventralizing action of Sonic hedgehog (Shh) which is mediated through the patterned expression of homeodomain (HD) and basic heli-loo-helix (bHLH) transcription factors.