EFFECT OF CALORIC VESTIBULAR STIMULATION ON MEMORY ASWATHY GOPINATH. MASTERS In MEDICAL PHYSIOLOGY. Under the guidance of. G SaiSailesh Kumar

Transcription

1 EFFECT OF CALORIC VESTIBULAR STIMULATION ON MEMORY BY ASWATHY GOPINATH Dissertation Submitted to the Kerala University of Health Sciences, Thrissur In partial fulfillment of the requirements for the degree of MASTERS In MEDICAL PHYSIOLOGY Under the guidance of G SaiSailesh Kumar DEPARTMENT OF PHYSIOLOGY LITTLE FLOWER INSTITUTE OF MEDICAL SCIENCES AND RESEARCH ANGAMALY 2016

2 KERALA UNIVERSITY OF HEALTH SCIENCES THRISSUR DECLARATION BY THE CANDIDATE I hereby declare that this dissertation entitled EFFECT OF CALORIC VESTIBULAR STIMULATION ON MEMORY is a bonafide and genuine research work carried out by me under theguidance of Mr.G SaiSailesh Kumar, Head of the Department, Department of Physiology, LIMSAR, Angamaly. Date: ASWATHY GOPINATH Place: Reg No:

3

4

5 COPYRIGHT Declaration by the Candidate I hereby declare that the Kerala University of Health Sciences, Thrissur shallhave the rights to preserve, use and disseminate this dissertation/thesis inprint or electronic format for academic/research purpose. Date: Place: ASWATHY GOPINATH

6 ABSTRACT OBJECTIVE This study was undertaken to provide an authoritative database for beneficial effects of vestibular stimulation and to suggest vestibular stimulation as a therapy for enhancement of cognition. MATERIALS AND METHOD Vestibular stimulation is performed by caloric vestibular stimulation. Scopolamine is used to induce partial amnesia. T-maze task is used to record acquisition and retention of the memory score before and after vestibular stimulation. RESULTS Memory scores were significantly different between scopolamine induced amnesia control and hot water vestibular stimulation groups. Memory scores between scopolamine induced amnesia cold and hot vestibular stimulation groups were highly significant. CONCLUSION This study categorically confirms that caloric vestibular stimulation with hot water enhances cognition. Hence this study certainly merits further studies with higher sample size to confirm whether caloric vestibular stimulation can be recommended for enhancement of cognition. KEYWORDS: Acquisition; Caloric vestibular stimulation; Cognition; Memory; Retention; Amnesia. I

7 TABLE OF CONTENTS SL.NO: CONTENTS PAGES 1. INTRODUCTION 1 2. OBJECTIVE 4 3. REVIEW OF LITERATURE 6 4. METHODOLOGY RESULTS DISCUSSION CONCLUSION REFERENCES ANNEXURES 66 II

8 LIST OF FIGURES SL.NO: FIGURES PAGES 1 Membranous labyrinth 8 2 Vestibular connections with central nervous 15 system 3 Subject receiving galvanic vestibular 17 stimulation 4 Memory systems that has been discovered in 23 the snail aplysia 5 T maze 35 6 Tmaze 38 7 Application of caloric vestibular stimulation 40 8 Acquisition memory scores 42 III

9 9 Acquisition memory scores Retention memory scores Retention memory scores 45 IV

10 ANNEXURES SL.NO: TITLE PAGES 1 Acknowledgement 67 2 List of abbreviations used 69 3 Publication 70 V

11 INTRODUCTION 1

12 CHAPTER I INTRODUCTION The vestibular apparatus is a membranous structure consisting of three semicircular canals connected at their base to the utricle, saccule and endolymphatic sac. The need of vestibular stimulation can be observed throughout the life. 1 Vestibular system is having extensive connections with hippocampus, raphe nucleus, locus ceruleus, thalamus, amygdala, insular cortex, anterior cingulated cortex, prefrontal cortex, cerebellum, occipital cortex, putamen, parietal lobe and other areas of brain which plays key role in cognitive processes. 2 Vestibular stimulation not only contributes for regulation of posture and equilibrium but also relieves stress, cancer pain, promotes sleep, improves immunity, improves cognition and also treats endocrine disorders. 3-5 Memory is the acquisition, storage and retrieval of sensory information. 6 The most common mazes used to study learning and memory are T-maze and R-maze. 7 Severe memory problems are observed in brain disorders like Alzheimer s disease, stroke, Parkinson s disease, Korsakoff syndrome, brain infections, brain tumors, seizures etc. Currently the drugs available for the treatment of cognitive disorders cannot cure the disorder, but only delay the loss of mental abilities or provide relief for short period of time. These drugs are expensive and have many side-effects. 8, 9 Hence there is need for a different approach to the treatment for cognitive disorders which is effective, 10, 11 affordable and with no or less side effects. 2

13 Patients with vestibular dysfunctions reported navigational and spatial memory impairments. 2 Vestibular system triggers a range of changes in cognition, emotion and personality through controlling autonomic functions. 2 Caloric vestibular stimulation is a safe non-invasive method of stimulating brain areas related to cognitive function. Clinically caloric vestibular stimulation is used as a diagnostic technique to investigate vestibular function. Caloric vestibular stimulation consists of a water irrigation of the external auditory canal, which induces a change in the temperature that leads to convection currents in the semicircular canals. This evokes a slow-phase nystagmus toward the stimulated ear and it elicits sensations of virtual body rotations and vertigo. 12 Caloric vestibular stimulation modulates sensory and cognitive functions in healthy participants and brain damaged patients. 13 Vestibular system can modulate cognition through hippocampus, through HPA axis through limbic system and neo cortex. 4 Vestibular stimulation activates areas of brain which are involved in learning and memory. 14 Caloric vestibular stimulation increases acetylcholine release from rat hippocampus and also enhances longterm potentiation via activation of cholinergic septo-hippocampal cells. 15,16 Vestibular dysfunction involves a complex syndrome characterized not only by reflex deficits but also by attention and memory deficits and anxiety disorders and depression. 17 3

14 OBJECTIVES 4

15 CHAPTER 2 OBJECTIVES MAIN To observe the effect of hot and cold water caloric vestibular stimulation on acquisition and retention. OTHER To compare the effect of caloric vestibular stimulation with hot water and cold water on acquisition and retention. HYPOTHESIS Caloric Vestibular stimulation may have positive impact on acquisition and retention and hot water caloric vestibular stimulation may be more beneficial than cold water caloric vestibular stimulation. 5

16 REVIEW OF LITERATURE 6

17 CHAPTER 3 REVIEW OF LITERATURE VESTIBULAR APPARATUS Vestibular system maintains the control of movement and sense of balance. 17 It is located in a bony chamber in the petrous portion of temporal bone called the bony labyrinth. Within this system are membranous tubes and chambers called the membranous labyrinth (Figure 1), which is the functional part. It is composed mainly of the cochlea (ductuscochlearis); three semicircular canals; and two large chambers, the utricle and saccule. "Maculae"-Sensory Organs of the Utricle and Saccule is Located on the inside surface of each utricle and saccule, measuring about 2mm in diameter. At the inferior surface of the utricle, macula lies in the horizontal plane and plays an important role in determining orientation of the head when the head is upright. Conversely, the macula of the saccule is located mainly in a vertical plane and signals head orientation when the person is lying down. Each macula is covered by a gelatinous layer in which many small calcium carbonate crystals called statoconia are embedded. In the macula there are thousands of hair cells. The cilia are projected into the gelatinous layer. The bases and sides of the hair cells synapse with sensory endings of the vestibular nerve. The calcified statoconia have a specific gravity two to three times the specific gravity of 7

18 surrounding fluid and tissues. The weight of the statoconia bends the cilia in the direction of gravitational pull. 18 FIGURE 1: MEMBRANOUS LABYRINTH 18 Directional Sensitivity of the Hair Cells-Kinocilium Each hair cell has 50 to 70 small cilia called stereocilia, plus one large cilium, the kinocilium, The kinocilium is always located to one side, and the stereocilia become progressively shorter toward the other side of the cell. Minute filamentous attachments connect the tip of each stereocilium to the next longer stereocilium and finally to the 8

19 kinocilium. Because of these attachments, when the stereocilia and kinocilium bend in the direction of the kinocilium, the filamentous attachments pulls stereocilia outward from the cell body. This opens several hundred fluid channels in the neuronal cell membrane around the bases of the stereocilia, as these can conduct positive ions, large number of positive ions flow from the endolymphatic fluid into the cell, causing receptor membrane depolarization. This causes bending of stereocilia to opposite direction which reduces the tension on attachment,this closes the ion channels, causing hyperpolarization Under normal resting conditions, the nerve fibers leading from the hair cells transmit continuous nerve impulses at a rate of about 100 per second. When the stereocilia are bent toward the kinocilium, the impulses are transmitted at a high rate. When the cilia bend away from the kinocilium the rate is decreased,oftenly turning it off completely. Therefore, as the orientation of the head in space changes, cilia are bend accordingly. Thus appropriate signals are transmitted to the brain to control equilibrium. 18 Semicircular Ducts The three semicircular ducts in each vestibular apparatus, known as the anterior, posterior, and lateral (horizontal) semicircular ducts, are arranged at right angles to one another so that they represent all three planes in space. Each semicircular duct has an enlargement at one of its ends called the ampulla, and the ducts and ampulla are filled with a fluid called endolymph. Flow of this fluid through one of the ducts and through its ampulla excites the sensory organ of the ampulla in the following manner. 9

20 In each ampulla a small crest can be seen called a crista ampullaris. On top of this crista is a loose gelatinous tissue mass, the cupula. When a person's head begins to rotate in any direction, the inertia of the fluid in one or more of the semicircular ducts causes the fluid to remain stationary while the semicircular duct rotates with the head. This causes fluid to flow from the duct and through the ampulla, bending the cupula to one side. Rotation of the head in the opposite direction causes the cupula to bend to the opposite side. Into the cupula are projected hundreds of cilia from hair cells located on the ampullary crest. The kinocilia of these hair cells are all oriented in the same direction in the cupula, and bending the cupula in that direction causes depolarization of the hair cells, whereas bending it in the opposite direction hyperpolarizes the cells. Then, from the hair cells, appropriate signals are sent by way of the vestibular nerve to apprise the central nervous system of a change in rotation of the head and the rate of change in each of the three planes of space. 18 Function of the Utricle and Saccule in the Maintenance of Static Equilibrium It is especially important that the hair cells are all oriented in different directions in the maculae of the utricles and saccules so that with different positions of the head, different hair cells become stimulated. The "patterns" of stimulation of the different hair cells appraise the brain of the position of the head with respect to the pull of gravity. In turn, the vestibular, cerebellar, and reticular motor nerve systems of the brain excite appropriate postural muscles to maintain proper equilibrium. This utricle and saccule system functions extremely effectively for maintaining equilibrium when 10

21 the head is in the near-vertical position. Indeed, a person can determine as little as half a degree of disequilibrium when the body leans from the precise upright position. 18 Detection of Linear Acceleration by the Utricle and Saccule Maculae When the body is suddenly thrust forward, that is, when the body accelerates-the statoconia, which have greater mass inertia than the surrounding fluid, fall backward on the hair cell cilia, and information of disequilibrium is sent into the nervous centres, causing the person to feel as though he or she were falling backward. This automatically causes the person to lean forward until the resulting anterior shift of the statoconia exactly equals the tendency for the statoconia to fall backward because of the acceleration. At this point, the nervous system senses a state of proper equilibrium and leans the body forward no further. Thus, the maculae operate to maintain equilibrium during linear acceleration in exactly the same manner as they operate during static equilibrium. The maculae do not operate for the detection of linear velocity. When runners first begin to run, they must lean far forward to keep from falling backward because of initial acceleration, but once they have achieved running speed, if they were running in a vacuum, they would not have to lean forward. When running in air, they lean forward to maintain equilibrium only because of air resistance against their bodies; in this instance, it is not the maculae that make them lean but air pressure acting on pressure end-organs in the skin, which initiate appropriate equilibrium adjustments to prevent falling

22 Detection of Head Rotation by the Semicircular Ducts When the head suddenly begins to rotate in any direction (called angular acceleration), the endolymph in the semicircular ducts, because of its inertia, tends to remain stationary while the semicircular ducts turn. This causes relative fluid flow in the ducts in the direction opposite to head rotation. A typical discharge signal from a single hair cell in the crista ampullaris when an animal is rotated for 40 seconds, occurs (1) Even when the cupula is in its resting position, the hair cell emits a tonic discharge of about 100 impulses per second. (2) When the animal begins to rotate, the hairs bend to one side and the rate of discharge increases greatly. (3) With continued rotation, the excess discharge of the hair cell gradually subsides back to the resting level during the next few seconds. The reason for this adaptation of the receptor is that within the first few seconds of rotation, back resistance to the flow of fluid in the semicircular duct and past the bent cupula causes the endolymph to begin rotating as rapidly as the semicircular canal itself; then, in another 5 to 20 seconds, the cupula slowly returns to its resting position in the middle of the ampulla because of its own elastic recoil. When the rotation suddenly stops, exactly opposite effects take place: The endolymph continues to rotate while the semicircular duct stops. This time, the cupula bends in the opposite direction, causing the hair cell to stop discharging entirely. After another few seconds, the endolymph stops moving and the cupula gradually returns to its resting position, thus allowing hair cell discharge to return to its normal tonic level. Thus, the 12

23 semicircular duct transmits a signal of one polarity when the head begins to rotate and of opposite polarity when it stops rotating. 18 "Predictive" Function of the Semicircular Duct System in the Maintenance of Equilibrium They detect is that the person's head is beginning or stopping to rotate in one direction or another. Therefore, the function of the semicircular ducts is not to maintain static equilibrium or to maintain equilibrium during steady directional or rotational movements. Yet loss of function of the semicircular ducts does cause a person to have poor equilibrium when attempting to perform rapid, intricate changing body movements. If a person is running forward rapidly and then suddenly begins to turn to one side, he or she will fall off balance a fraction of a second later unless appropriate corrections are made ahead of time. But the maculae of the utricle and saccule cannot detect that he or she is off balance until after this has occurred. The semicircular ducts, however, will have already detected that the person is turning, and this information can easily appraise the central nervous system of the fact that the person will fall off balance within the next fraction of a second or so unless some anticipatory correction is made. In other words, the semicircular duct mechanism predicts that disequilibrium is going to occur and thereby causes the equilibrium centers to make appropriate anticipatory preventive adjustments. This helps the person maintain balance before the situation can be corrected. Removal of the flocculonodular lobes of the cerebellum prevents normal detection of semicircular duct signals but has less effect on detecting 13

24 macular signals. It is especially interesting that the cerebellum serves as a "predictive" organ for most rapid movements of the body, as well as for those having to do with equilibrium. 18 Neuronal Connections of the Vestibular Apparatus with the Central Nervous System Most of the vestibular nerve fibers terminate in the brain stem in the vestibular nuclei, which are located approximately at the junction of the medulla and the pons. Some fibers pass directly to the brain stem reticular nuclei without synapsing and also to the Cerebellar Fastigial, Uvular, and Flocculonodular lobe nuclei (Figure 2). The fibers that end in the brain stem vestibular nuclei synapse with second-order neurons that also send fibers into the cerebellum, the vestibulospinal tracts, the medial longitudinal fasciculus, and other areas of the brain stem, particularly the reticular nuclei. The primary pathway for the equilibrium reflexes begins in the vestibular nerves, where the nerves are excited by the vestibular apparatus. The pathway then passes to the vestibular nuclei and cerebellum. Next, signals are sent into the reticular nuclei of the brain stem, as well as down the spinal cord by way of the vestibulospinal and reticulospinal tracts. The signals to the cord control the interplay between facilitation and inhibition of the many antigravity muscles, thus automatically controlling equilibrium. The flocculonodular lobes of the cerebellum are especially concerned with dynamic equilibrium signals from the semicircular ducts. In fact, destruction of these lobes results in almost exactly the same clinical symptoms as destruction of the semicircular 14

25 ducts themselves. That is, severe injury to either the lobes or the ducts causes loss of dynamic equilibrium during rapid changes in direction of motion but does not seriously disturb equilibrium under static conditions. It is believed that the uvula of the cerebellum plays a similar important role in static equilibrium. 18 FIGURE 2: VESTIBULAR CONNECTIONS WITH CENTRAL NERVOUS SYSTEM 18 Signals transmitted upward in the brain stem from both the vestibular nuclei and the cerebellum by way of the medial longitudinal fasciculus cause corrective movements of the eyes every time the head rotates, so the eyes remain fixed on a specific visual object. Signals also pass upward (either through this same tract or through reticular tracts) to the cerebral cortex, terminating in a primary cortical center for equilibrium located in the parietal lobe deep in the sylvian fissure on the opposite side of the 15

26 fissure from the auditory area of the superior temporal gyrus. These signals appraise the psyche of the equilibrium status of the body. 18 METHODS TO STIMULATE VESTIBULAR SYSTEM Motion devices Lectos pensiles (hanging bed), lecti suspense motus (floating beds) were used in setting body in motion to cure madness by Greek and Roman physicians. Later rotating chairs and rotating beds came into practice for the treatment of patients in asylum. The use of rotating chair also paved the way for stimulation of vestibular system and associated eye movements. This lead to the use of rotating chair as the diagnostic tool for the vestibular disorders. From 1920s large scale centrifuges were made to stimulate an increase of gravitational forces and study its effect on human physiology and cognition. 100 Caloric vestibular stimulation 62, 84 This method is done by irrigating external ear. Galvanic vestibular stimulation In this method vestibular nerve is activated through the skin over mastoid process. 74 Both otolith organs and semicircular ducts are stimulated in GVS. 75 Though the vestibular nerve is stimulated mainly the otolith organs are activated in GVS. 76 Electrical impulses are stimulating a viscous fluid that activates hair cells located in the inner ear, which are responsible for vertical orientation and linear movement (Figure 3). Rotational movement is detected by the semicircular canals, which are filled with a fluid called endolymph. It is important to avoid connecting the 16

27 electrodes to hair, because they are an insulator for electrical impulses that are sent to the nerve in the vestibular system. 77 Rotatory Vestibular Stimulation In this method the animal is rotated in cages at a particular frequency. In earlier works the speed of rotation was fixed at 100 revolutions per minute by trial and error method. 73 FIGURE 3: SUBJECT RECEIVING GALVANIC VESTIBULAR STIMULATION. 77 THOUGHTS, CONSCIOUSNESS, AND MEMORY Thought certainly involves simultaneous signals in many portions of the cerebral cortex, thalamus, limbic system, and reticular formation of the brain stem.we might formulate a provisional definition of a thought in terms of neural activity as follows: A thought results from a "pattern" of stimulation of many parts of the nervous system at 17

28 the same time, probably involving most importantly the cerebral cortex, thalamus, limbic system, and upper reticular formation of the brain stem. This is called the holistic theory of thoughts. The stimulated areas of the limbic system, thalamus, and reticular formation are believed to determine the general nature of the thought, giving it such qualities as pleasure, displeasure, pain, comfort, crude modalities of sensation, localization to gross areas of the body, and other general characteristics. However, specific stimulated areas of the cerebral cortex determine discrete characteristics of the thought, such as (1) Specific localization of sensations on the surface of the body and of objects in the fields of vision, (2) The feeling of the texture of silk, (3) Visual recognition of the rectangular pattern of a concrete block wall, (4) Other individual characteristics that enter into one's overall awareness of a particular instant. Consciousness can perhaps be described as our continuing stream of awareness of either our surroundings or our sequential thoughts. 18 Memory-Roles of Synaptic Facilitation and Synaptic Inhibition Memories are stored in the brain by changing the basic sensitivity of synaptic transmission between neurons as a result of previous neural activity. The new or facilitated pathways are called memory traces. They are important because once the traces are established; they can be selectively activated by the thinking mind to reproduce the memories. 18

29 Experiments in lower animals have demonstrated that memory traces can occur at all levels of the nervous system. Even spinal cord reflexes can change at least slightly in response to repetitive cord activation, and these reflex changes are part of the memory process. Also, long-term memories result from changed synaptic conduction in lower brain centers. However, most memory that we associate with intellectual processes is based on memory traces in the cerebral cortex. Positive and Negative Memory-"Sensitization" or "Habituation" of Synaptic Transmission Although we often think of memories as being positive recollections of previous thoughts or experiences, probably the greater share of our memories is negative, not positive. That is, our brain is inundated with sensory information from all our senses. If our minds attempted to remember all this information, the memory capacity of the brain would be rapidly exceeded. Fortunately, the brain has the capability to learn to ignore information that is of no consequence. This results from inhibition of the synaptic pathways for this type of information; the resulting effect is called habituation. This is a type of negative memory. Conversely, for incoming information that causes important consequences such as pain or pleasure, the brain has a different automatic capability of enhancing and storing the memory traces. This is positive memory. It results from facilitation of the synaptic pathways, and the process is called memory sensitization

30 CLASSIFICATION OF MEMORIES We know that some memories last for only a few seconds, whereas others last for hours, days, months, or years. For the purpose of discussing these, let us use a common classification of memories that divides memories into (1) Short-term memory, which includes memories that last for seconds or at most minutes unless they are converted into longer-term memories, (2) Intermediate long-term memories, which last for days to weeks but then fade away, (3) Long-term memory, which, once stored, can be recalled up to years or even a lifetime later. In addition to this general classification of memories, another type of memory, can be called working memory," which includes mainly short-term memory that is used during the course of intellectual reasoning but is terminated as each stage of the problem is resolved. 18 Memories are frequently classified according to the type of information that is stored. One of these classifications divides memory into declarative memory and skill memory, as follows: 1. Declarative memory basically means memory of the various details of an integrated thought, such as memory of an important experience that includes (1) memory of the surroundings, (2) memory of time relationships, (3) memory of causes of the experience, (4) memory of the meaning of the experience, and (5) memory of one's deductions that were left in the person's mind

31 2. Skill memory is frequently associated with motor activities of the person's body, such as all the skills developed for hitting a tennis ball, including automatic memories to (1) sight the ball, (2) calculate the relationship and speed of the ball to the racquet, and (3) deduce rapidly the motions of the body, the arms, and the racquet required to hit the ball as desired-all of these activated instantly based on previous learning of the game of tennis-then moving on to the next stroke of the game while forgetting the details of the previous stroke. 18 Short-Term Memory Short-term memory is typified by one's memory of 7 to 10 numerals in a telephone number (or 7 to 10 other discrete facts) for a few seconds to a few minutes at a time but lasting only as long as the person continues to think about the numbers or facts. Many physiologists have suggested that this short-term memory is caused by continual neural activity resulting from nerve signals that travel around and around a temporary memory trace in a circuit of reverberating neurons. It has not yet been possible to prove this theory. Another possible explanation of short-term memory is presynaptic facilitation or inhibition. This occurs at synapses that lie on terminal nerve fibrils immediately before these fibrils synapse with a subsequent neuron. The neurotransmitter chemicals secreted at such terminals frequently cause facilitation or inhibition lasting for seconds up to several minutes. Circuits of this type could lead to short-term memory

32 Intermediate Long-Term Memory Intermediate long-term memories may last for many minutes or even weeks. They will eventually be lost unless the memory traces are activated enough to become more permanent; then they are classified as long-term memories. Experiments in primitive animals have demonstrated that memories of the intermediate long-term type can result from temporary chemical or physical changes, or both, in either the synapse presynaptic terminals or the synapse postsynaptic membrane, changes that can persist for a few minutes up to several weeks. These mechanisms are so important that they deserve special description. 18 Memory Based on Chemical Changes in the Presynaptic Terminal or Postsynaptic Neuronal Membrane There are two synaptic terminals. One terminal is from a sensory input neuron and terminates directly on the surface of the neuron that is to be stimulated; this is called the sensory terminal. The other terminal is a presynaptic ending that lies on the surface of the sensory terminal, and it is called the facilitator terminal (Figure 4). When the sensory terminal is stimulated repeatedly but without stimulation of the facilitator terminal, signals transmission at first is great, but it becomes less and less intense with repeated stimulation until transmission almost ceases. This phenomenon is habituation. It is a type of negative memory that causes the neuronal circuit to lose its response to repeated events that are insignificant. Conversely, if a noxious stimulus excites the facilitator terminal at the same time that the sensory terminal is stimulated, then instead of the transmitted signal into the 22

33 postsynaptic neuron becoming progressively weaker, the ease of transmission becomes stronger and stronger; and it will remain strong for minutes, hours, days, or, with more intense training, up to about 3 weeks even without further stimulation of the facilitator terminal. Thus, the noxious stimulus causes the memory pathway through the sensory terminal to become facilitated for days or weeks thereafter. It is especially interesting that even after habituation has occurred, this pathway can be converted back to a facilitated pathway with only a few noxious stimuli. 18 FIGURE 4: MEMORY SYSTEMS THAT HAS BEEN DISCOVERED IN THE SNAIL APLYSIA

34 Molecular Mechanism of Intermediate Memory Mechanism for Habituation At the molecular level, the habituation effect in the sensory terminal results from progressive closure of calcium channels through the terminal membrane, though the cause of this calcium channel closure is not fully known. Nevertheless, much smaller than normal amounts of calcium ions can diffuse into the habituated terminal, and much less sensory terminal transmitter is therefore released because calcium entry is the principal stimulus for transmitter release. 18 Mechanism for Facilitation In the case of facilitation, at least part of the molecular mechanism is believed to be the following: 1. Stimulation of the facilitator presynaptic terminal at the same time that the sensory terminal is stimulated causes serotonin release at the facilitator synapse on the surface of the sensory terminal. 2. The serotonin acts on serotonin receptors in the sensory terminal membrane, and these receptors activate the enzyme adenyl cyclase inside the membrane. The adenyl cyclase then causes formation of cyclic adenosine monophosphate (camp) also inside the sensory presynaptic terminal. 3. The cyclic AMP activates a protein kinase that causes phosphorylation of a protein that itself is part of the potassium channels in the sensory synaptic terminal membrane; this in turn blocks the channels for potassium conductance. The blockage can last for minutes up to several weeks. 24

35 4. Lack of potassium conductance causes a greatly prolonged action potential in the synaptic terminal because flow of potassium ions out of the terminal is necessary for rapid recovery from the action potential. 5. The prolonged action potential causes prolonged activation of the calcium channels, allowing tremendous quantities of calcium ions to enter the sensory synaptic terminal. These calcium ions cause greatly increased transmitter release by the synapse, thereby markedly facilitating synaptic transmission to the subsequent neuron. Thus, in a very indirect way, the associative effect of stimulating the facilitator terminal at the same time that the sensory terminal is stimulated causes prolonged increase in excitatory sensitivity of the sensory terminal, and this establishes the memory trace. 18 Long-Term Memory There is no obvious demarcation between the more prolonged types of intermediate long-term memory and true long-term memory. The distinction is one of degree. However, long-term memory is generally believed to result from actual structural changes, instead of only chemical changes, at the synapses, and these enhance or suppress signal conduction. 18 Structural Changes Occur in Synapses during the Development of Long-Term Memory Electron microscopic pictures taken from invertebrate animals have demonstrated multiple physical structural changes in many synapses during development of long- 25

36 term memory traces. The structural changes will not occur if a drug is given that blocks DNA stimulation of protein replication in the presynaptic neuron; nor will the permanent memory trace develop. Therefore, it appears that development of true longterm memory depends on physically restructuring the synapses themselves in a way that changes their sensitivity for transmitting nervous signals. The most important of the physical structural changes that occur are the following: 1. Increase in vesicle release sites for secretion of transmitter substance, 2. Increase in number of transmitter vesicles released, 3. Increase in number of presynaptic terminals, 4. Changes in structures of the dendritic spines that permit transmission of stronger signals, Thus, in several different ways, the structural capability of synapses to transmit signals appears to increase during establishment of true long-term memory traces. 18 Number of Neurons and Their Connectivities Often Change Significantly During Learning During the first few weeks, months, and perhaps even year or so of life, many parts of the brain produce a great excess of neurons and the neurons send out numerous axon branches to make connections with other neurons. If the new axons fail to connect with appropriate neurons, muscle cells, or gland cells, the new axons themselves will dissolute within a few weeks. Thus, the number of neuronal connections is determined by specific nerve growth factors released retrogradely from the stimulated cells. 26

37 Furthermore, when insufficient connectivity occurs, the entire neuron that is sending out the axon branches might eventually disappear. 18 Therefore, soon after birth, there is a principle of "use it or lose it" that governs the final number of neurons and their connectivities in respective parts of the human nervous system. This is a type of learning. For example, if one eye of a newborn animal is covered for many weeks after birth, neurons in alternate stripes of the cerebral visual cortex-neurons normally connected to the covered eye-will degenerate, and the covered eye will remain either partially or totally blind for the remainder of life. Until recently, it was believed that very little learning" is achieved in adult human beings and animals by modification of numbers of neurons in the memory circuits; however, recent research suggests that even adults use this mechanism to at least some extent. 18 CONSOLIDATION OF MEMORY For short-term memory to be converted into long-term memory that can be recalled weeks or years later, it must become "consolidated." That is, the short-term memory if activated repeatedly will initiate chemical, physical, and anatomical changes in the synapses that are responsible for the long-term type of memory. This process requires 5 to 10 minutes for minimal consolidation and 1 hour or more for strong consolidation. For instance, if a strong sensory impression is made on the brain but is then followed within a minute or so by an electrically induced brain convulsion, the sensory experience will not be remembered. Likewise, brain concussion, sudden 27

38 application of deep general anesthesia, or any other effect that temporarily blocks the dynamic function of the brain can prevent consolidation. Consolidation and the time required for it to occur can probably be explained by the phenomenon of rehearsal of the short-term memory as follows: Rehearsal Enhances the Transference of Short-Term Memory into Long-Term Memory Studies have shown that rehearsal of the same information again and again in the mind accelerates and potentiates the degree of transfer of short-term memory into long-term memory and therefore accelerates and enhances consolidation. The brain has a natural tendency to rehearse newfound information, especially newfound information that catches the mind's attention. Therefore, over a period of time, the important features of sensory experiences become progressively more and more fixed in the memory stores. This explains why a person can remember small amounts of information studied in depth far better than large amounts of information studied only superficially. It also explains why a person who is wide awake can consolidate memories far better than a person who is in a state of mental fatigue. 18 New Memories Are Codified During Consolidation One of the most important features of consolidation is that new memories are codified into different classes of information. During this process, similar types of information are pulled from the memory storage bins and used to help process the new information. The new and old are compared for similarities and differences, and part of the storage process is to store the information about these similarities and 28

39 differences, rather than to store the new information unprocessed. Thus, during consolidation, the new memories are not stored randomly in the brain but are stored in direct association with other memories of the same type. This is necessary if one is to be able to "search" the memory store at a later date to find the required information. 18 Role of Specific Parts of the Brain in the Memory Process Hippocampus Promotes Storage of Memories-Anterograde Amnesia After Hippocampal Lesions The hippocampus is the most medial portion of the temporal lobe cortex, where it folds first medially underneath the brain and then upward into the lower, inside surface of the lateral ventricle. The two hippocampi have been removed for the treatment of epilepsy in a few patients. This procedure does not seriously affect the person's memory for information stored in the brain before removal of the hippocampi. However, after removal, these people have virtually no capability thereafter for storing verbal and symbolic types of memories (declarative types of memory) in long-term memory, or even in intermediate memory lasting longer than a few minutes. Therefore, these people are unable to establish new long-term memories of those types of information that are the basis of intelligence. This is called anterograde amnesia. The hippocampi are among the most important output pathways from the "reward" and "punishment" areas of the limbic system. Sensory stimuli or thoughts that cause pain or aversion excite the limbic punishment centers, and stimuli that cause pleasure, background mood and motivations of the person. Among these motivations is the drive in the brain to remember those experiences and thoughts that are either pleasant or unpleasant. The hippocampus especially and to a lesser degree 29

40 the dorsal medial nuclei of the thalamus, another limbic structure, have proved especially important in making the decision about which of our thoughts are important enough on a basis of reward or punishment to be worthy of memory. 18 Retrograde Amnesia-Inability to Recall Memories from the Past When retrograde amnesia occurs, the degree of amnesia for recent events is likely to be much greater than for events of the distant past. The reason for this difference is probably that the distant memories have been rehearsed so many times that the memory traces are deeply ingrained, and elements of these memories are stored in widespread areas of the brain. In some people who have hippocampal lesions, some degree of retrograde amnesia occurs along with anterograde amnesia, which suggests that these two types of amnesia are at least partially related and that hippocampal lesions can cause both. However, damage in some thalamic areas may lead specifically to retrograde amnesia without causing significant anterograde amnesia. A possible explanation of this is that the thalamus may play a role in helping the person "search" the memory storehouses and thus "read out" the memories. That is, the memory process not only requires the storing of memories but also an ability to search and find the memory at a later date. 18 Hippocampi Are Not Important in Reflexive Learning People with hippocampal lesions usually do not have difficulty in learning physical skills that do not involve verbalization or symbolic types of intelligence. For instance, these people can still learn the rapid hand and physical skills required in many types of sports. This type of learning is called skill learning or reflexive learning; it depends on 30

41 physically repeating the required tasks over and over again, rather than on symbolical rehearsing in the mind 18. Vestibular system is having connections with hypothalamic nuclei, autonomic system, dorsal and median raphe nuclei, substantia nigra, hippocampal formation and controls pancreatic secretion. 19 VESTIBULAR SYSTEM AND COGNITION Cognition means thinking and awareness. 20 It is the process by which sensory inputs are processed and reused as per need.it is seen that vestibular dysfunction causes memory and anxiety disorders 21.Lesion of vestibular system causes defect in attention, learning, short term and working memory There are four major pathways put forward for transmissions of vestibular information to cortex: (a) Vestibular-thalamo-cortical pathway: It transmits spatial information via parietal cortex to the hippocampus. (b) Transmits information about direction of head from dorsal tegmental nucleus via lateral mammilary nucleus and anterodorsal nucleus of the thalamus to the cortex. (c) Transmits memory via the Nucleus reticularis, Pontis oralis, Supramammillary nucleus to the hippocampus. (d) Transmits information about spatial learning via cerebellum and ventral lateral nucleus of the thalamus

42 Vestibular-thalamo-cortical pathway Medial longitudinal fasciculus: It connects superior vestibular nucleus to Ipsilateral central lateral, Bilateral ventro-postero-lateral and Ventro lateral thalamic nuclei.medial vestibular nucleus to contralateral central lateral and Bilateral ventropostero-lateral thalamic nuclei. Descending vestibular nuclei to medial geniculate nucleus of opposite side Ascending tract of Deiter: Superior and medial vestibular nuclei to central lateral,ventral-posterior-lateral and ventral-lateral thalamic nuclei Ipsilateral vestibulo-thalamic tract: It transmits vestibular information to thalamus about otolithic signals. 48 The pathway from the dorsal tegmental nucleus to the entorhinal cortex Head directional signal is transmitted from vestibular nuclei to tegmental nucleus then lateral mammillary nucleus,anterodorsal nucleus of thalamus,posterior subiculum,parasubiculum and medial entorhinal cortex. 49 The pathway from the nucleus reticularis pontis oralis,to the medial septum to the hippocampus Ascending signals activates supramammillary nucleus which convert it to pattern of rhythm. This pattern activates hippocampal interneurons and principal cells. 50 The pathway from cerebellum to parietal cortex Cerebellum is connected via rostral vermis,fastigial nucleus,intervening midline folia to hippocampus which is a direct connection Thalamic connection is proposed via the fastigial,dentate and interposed nuclei to the thalamus

43 57, 58 Electrical stimulation of vestibular system elicit response in caudate nucleus. Literature suggests that vestibular stimulation have a positive effect on cognition especially on spatial tasks through otolithic and visual stimulation and it is independent of age. 27 Galvanic vestibular stimulation have a positive effect on perspective-taking tasks. 28 Vestibular stimulation proves to be a better option for attention-deficit/hyperactivity disorder. 29 Acetylcholine can facilitate long term potentiation in hippocampus and modulate learning and memory through induced vestibular stimulation. 30 Cognition can be affected by chronic stress by the action of stress related changes in the body. 31 Vestibular stimulation inhibit both HPA and SAM axes which in turn decrease cortisol level and inhibiting the stress related change in body. 32 Neocortex and limbic system are also involved in cognition. 33 Caloric vestibular stimulation link vestibular system and limbic system effecting cognition. 34 There are nine major vestibuar cortical areas which have a important role in spatial coginition. 35 Parieto-insular vestibular cortex and temporal junction Most of the neurons in parieto-insular vestibular cortex (PIVSC) and temporal junction respond to the vestibular stimulation. So PIVSC is called the principal cortex. 36 Anterior parietal cortex Somatosensory information from head,neck,upper limbs and the vestibular input are integrated in anterior parietal cortex

44 This plays a major function in differentiating self from object motion. 35 Posterior parietal and medial superior temporal cortices Parietal cortex functions as a center for spatial representation and provides information about movement. 37 Medial superior temporal cortex encodes self motion information from vestibular and visual inputs and compare it with object motion. It also provides details about spatial orientation. 35 Cingulate gyrus and retrosplenial cortex Anterior and posterior cingulated gyrus are activated in caloric vestibular stimulation. 38 Retrospinal cortex encodes information about navigation and path integration. 39,40 Hippocampal and parahippocampal cortices These play a major role in spatial coginition. 41 Bilateral vestibular dysfunction cause bilateral atrophy of hippocampus and memory deficit. 42 CALORIC VESTIBULAR STIMULATION Caloric stimulation for vestibular diagnostic test was developed by Barany Caloric stimulation is done by irrigating external ear with warm or cold water. This causes movement in semicircular canals which in turn signals vestibular nuclei by vestibular nerve and activation of contralateral cortical and subcortical structures. Subjects experience vertigo after seconds. 62 CVS when given to phantom limb subjects, reported significant relive of pain. 63 Reduction of pain was also observed when CVS was administered to thalamic syndrome subjects. 64 and in complex regional pain syndrome. 65 Left ear stimulation 34

45 improved elevated mood by 20 in young mania rating scale. 66 CVS of left ear improves spatial memory and that of right ear improves verbal memory. 67 T MAZE T-maze is a hollow wooden box in the shape of letter T. This maze is commonly used for behavioral studies (Figure 5). It can be used for various cognitive tests such as reference memory and working memory. 68 T maze is also used for assessing behavioral flexibility. Left and right discrimination task is mainly used to analyze rodents reference memory. 69 In the present study the protocol followed is in relation with learning of T maze. 70 In a recent study, tested the duration of memory on the T- maze. Ten rats were tested on a T-maze with delays up to 10 min. A delay-dependent decrease in memory was observed, with above chance performance at 5 min, but performance at a chance level with 10 min. 71 Nowadays automated mazes are used for short term memory test which are more reliable than standard T maze. 72 Figure 5: T MAZE 72 35

46 RESEARCH METHODOLOGY 36

47 CHAPTER 4 RESEARCH METHODOLOGY RESEARCH APPROACH The present study was an experimental study conducted between December February VARIABLES Independent variable- Caloric vestibular stimulation Dependent variable- Acquisition and Retention ETHICAL CONSIDERATION The present study was approved by institutional animal ethical committee of Little Flower Institute of Medical Science and Research. ( , No EC/6). MATERIALS AND METHODS Animals: 18 healthy, adult male albino rats of Wistar strain with body weight ranging between (50-125g) were used in the present study. Rats were housed in polypropylene cages (30x22x14 cm), fed with standard-chow and water ad libitum. Rats are randomly assigned into three groups: Group A: (n=6) Control group-partial amnesia was induced by administration of scopolamine and no vestibular stimulation. Group B: (n=6) Partial amnesia was induced by administration of scopolamine and provided caloric vestibular stimulation with cold water for 30 days. 37

48 Group C: (n=6) Partial amnesia was induced by administration of scopolamine and provided caloric vestibular stimulation with hot water for 30 days. 1. INCLUSION CRITERIA Healthy adult rats Male rats 2. EXCLUSION CRITERIA Female rats Unhealthy diseased rats MATERIALS T-maze: (Figure-6) The T-maze is made of wood with smooth polished surface. It consists of a stem (35 x 12 cm), a choice area (12 x 12 cm) and two arms (35 x 12 cm); at the end of each arm contain a food well. The sidewalls are 40 cm high. The choice area is separated from the arms by a sliding door. 6 FIGURE 6: T-MAZE 38

49 T-maze task In the orientation phase, the starved rats were allowed to spend 10 minutes / day for three days in the T-maze and trained to collect food pellet from the food wells. During the acquisition test, all the rats were given six trials / day with an inter trial interval of one hour. Each trial consists of four sample and choice run. In the sample run, the rat was placed at the start end of the T-maze stem. Allowed to move towards one arm and collect the food pellet, while keeping the sliding door of other arm closed. In the choice run, the rat was placed at the start end of stem and both arms were kept open. If the rat visits the same arm as that of sample run, it was recorded as correct score and the rat was rewarded with food. Instead, if the rat visits the alternate arm, it was recorded as error and the rat was not allowed to eat food pellet. There was an interval of 30s between each run. After 3 days of orientation phase behavioral task was performed. This task was continued till we get the full score. Once the full score is recorded, ten days gap was given. After ten days retention test was conducted and memory score was recorded. From the next day onwards scopolamine was administered intraperitoneal for 9 days at 10am daily. From 10 th day, cold and hot caloric vestibular stimulation was given for 30 days to group B and group C respectively. From 31 st day behavioral task was conducted as explained earlier Drugs: Buscopan tablets manufactured by Cadila Healthcare limited, is used in the present study. Each Buscopan tablet contained Hyoscine (scopolamine) Butylbromide I P 10 mg and excipients (q. s.). The tablets were powdered and mixed with 50ml 39

50 sterile 0.9% w/v normal saline. It was administered by intraperitoneal injection at a dose of 1 mg / Kg. Scopolamine was injected at a dose of 1mg/Kg body weight of rat. Caloric vestibular stimulation: (Figure 7) The middle ear cavity was irrigated with hot (44 degree centigrade) or cold (30 degree centigrade) (Figure - 2) water through a 62, 84 polyethylene tube for 30 days. FIGURE 7: APPLICATION OF CALORIC VESTIBULAR STIMULATION ANALYSIS AND INTERPRETATION Data Analysis Data was analyzed by SPSS 20.0 by Two Way ANOVA and further conclusions were drawn by bonferroni test. 40