BEHAVIOR OF THE SQUARE WAVE IN BIOLOGICAL STRUCTURES

This study is to experimentally investigate the behavior of quadratic waves in biological structures. Transcutaneous frequency stimulation was regulated so that the system provided high frequency and low electrical voltage to observe possible changes in deeper layers of the anatomical structure analyzed. Electrodes were positioned from the dermis and probing electrodes were positioned at the subdermal, intradermal, skeletal muscle, periosteum, endosteum, and medulla levels. A function generator connected to a digital oscilloscope produced a square wave of 33 kHz at 10µs pulse speed and voltage of 16 volts DC. A voltage decay was observed as the signal reached deeper layers of different tissues. However, the frequency of 33 kHz remained uniform even at the bone marrow level. This observation opens the way for the application of the square electronic frequency as a stimulus for drug activation.


INTRODUCTION
We are fundamentally electronic beings. Our cells use ionic currents and electric fields to drive the transduction mechanism. Among numerous examples, it can be mentioned rhodopsin, a protein pigment found in retinal rods important for vision, which is activated by light that triggers the cascade of amplification signals that leads to a reduction in the cGMP gating of the Na+ channel. The change in the ionic current modifies the membrane potential of the rod and influences signaling to other cells in the visual cortex that eventually form an image in our brains. Another example is hearing. Air compressions impinge on the eardrum. The neuromotor ciliary kinetics of inner ear cells triggers the opening of ion channels for transduction of vibration into membrane potential, which converts into electrical signals that the brain interprets as noise. In simple terms, cells, tissues, and organs constantly send out electronic signals to maintain homeostasis, enable physiological functions, and respond to injury and infection. At the slightest signaling error, a multitude of pathologies can occur. The knowledge of electronic aspects of the human body leads to the search for methods that can help the correction of organic dysfunctions (1) (2) .
Technically all organs and systems are regulated by the brain. Neural circuits communicate through electronic triggers at certain frequencies and wavelengths. Even the endocrine system is under the control of the central nervous system by a complex set of feedback mechanisms. Most drugs interact electronically with final (neural) or endocrine receptors. Therefore, Electroceuticals appears as a technology that aims to complement and eventually replace some drugs of a chemical nature with electronic stimuli (digital drugs) capable of creating a response in cellular ultrastructures (2) (4) .

DEFINITION OF ELECTROCEUTICALS
It is a recently coined term for an ancient therapeutic modality that encompasses all of bioelectronic medicine. Basically, this refers to the electrical stimulation that can affect and/or modify body functions through in situ neural implants, such as cochlear implants, retinal implants or spinal cord stimulators for pain relief, but also cardiac pacemakers and implantable defibrillators. Recently, the field has expanded to include deep brain stimulation and electrical vagus nerve stimulation. Also called bioelectronics, the idea is that small electronic implants are capable of treating a wide range of chronic diseases, such as diabetes, asthma, airway obstruction, arthritis, hypertension and heart disease, as well as gastrointestinal problems (5) .

MECHANISM OF ACTION
Briefly, the mechanism of action of electroceuticals is based on electrostimulation, whose access route is the vagus nerve (implant technique) (3) .
This nerve is long and originates from the skull, precisely located behind the medulla oblongata -the structure that connects the brain with the spinal cord -it passes through the jugular foramen, the neck, and thorax to the stomach. The first step in the development of this therapy is to accurately map the neural circuits associated with the disease and consequently its treatment (5) .

ELECTRONIC INTERFERENCE
The existence of free ions, dipoles, and polarized molecules within the human body allows cells, tissues, and organs to generate and conduct electricity. Bioelectric voltage plays a vital role in regulating physiological processes. Based on this and on the complexity of vagal innervation, added to the production of electrical triggers from the neural network, sending electronic signals becomes a challenge, as it is assumed that such signals of endogenous and cellular nature flow through the body and generate a noisy electronic network difficult to control. A therapeutic electrical stimulus, for example, could suffer interference due to the organic electrical network itself, making the maneuver not fully effective (2) .

MAIN GOAL
In situ implants for electrostimulation have become unquestionably indispensable in modern health. Functionality, miniaturization, biocompatibility, and/or biodegradability are the main engineering focuses for the development and clinical application of these respective implants. This technology is focused mainly on developing wearable/implantable energy devices and advanced conformable electrodes for efficient stimulation of targets such as organs and tissues. Electrodes deliver electrical signals to the target tissue/organ (4) .

SPECIFIC OBJECTIVE
To present a possible alternative, non-invasive and economical evidence of using electronic frequency in the form of a square wave to stimulate the biological system.

Biological material probed
The experiment used the leg of a galliform bird of the species Gallus domesticus, collected from a slaughterhouse.
First, the ECRs were inserted aiming at contact with the respective histological areas to be studied: subdermis, intradermis, skeletal muscle, periosteum, endosteum, and medulla. Then, the dermis of the anatomical structure was cleaned with alcohol for EOI positioning. The fixation was done through a strip of insulating polymer based on cis-polyisoprene.  ECR electrode inserted in the bone marrow region.

Signal regulation
The equipment was adjusted to the following parameters: • Square wave

RESULTS
Immediately after triggering the incident wave, the data collected by the resulting pickup electrodes (ECR) showed voltage decay:     The square wave acts as an agitator of nanometric structures. In this case, biological tissues are naturally overdamped. The elastic reactance of the cell membrane is an example. The waveform may never actually reach the theoretical high and low levels, and if the system is underdamped, it will oscillate between high and low levels before stabilizing (9) .
A priori, according to graph 2, it can be deduced that the frequency stabilized at 33 kHz would represent that biological structures are invisible to this wave. However, by converting the respective frequency into nanometers, the effect reaches structures in the order of 7.1 nm. The result is in nanometers = 7.1, that is, the frequency of 33 kHz is capable of resonating with structures in this dimension. It is important to note that resonance refers to the vibrating system that induces another system to oscillate. On the scale of 7.1 nanometers, this would equate to the cell membrane thickness of bacteria, algae, fungi, protozoa, and animals. As the frequency increases, the nanometer scale decreases. Consequently, even smaller structures can be achieved.
Thus, it is possible to speculate that the application of ultrashort pulses (hammer effect), characteristic only of the quadratic wave, may cause reactions not only at the membrane level but also in any other nanostructure, such as ion channels of the order of 5 nm, which could be resonated at a frequency of 47 kHz.

CONCLUSION
The experiment indicates that kilohertz pulses can reach cellular structures and their external and internal substructures and therefore, possibly alter their functions. This is independent of the applied voltage. Even molecules could be stimulated. In this case, the frequency would be the acting agent.
Since the typical loading time constants of cells with dimensions of 10 μm are on the order of one hundred nanoseconds, submicrosecond electrical pulses are expected to cause different effects. New experiments must be conducted to evaluate the respective behaviors at the nanometer/micrometer level and, consequently, at the tissue level as a whole (6) .
Speculatively, it can be considered the application of the administration of pulse electric fields to activate molecules with therapeutic or prophylactic potential inside cells. Activation would take place through specific frequencies. In the case of chemotherapeutic agents, the influx or efflux of the drug through the receptors could be controlled frequently. The use of pulses would assist in this process, giving the agent a more direct path to the cytosol. Pulsatile electronic waves would allow access to the cytoplasmic target for drugs that could only cross the cytosol very poorly. It is important to emphasize that the frequency used is not ionizing, since it would have to be in the order of more than 20 MHz (7) .
There are some research possibilities to be developed in the future: ✓ Use of electronic frequency to activate a drug; ✓ Acting as a cell stimulator to accelerate tissue regeneration processes; ✓ Activation of defective cell receptors by frequency resonance; ✓ Adjuvant in antibiotic treatments.