Induction of hydrocephalus and its relation to Cell death, axonal damage, and cell birth in the immature rat brain

1. Induction of hydrocephalusand its relation to Cell death, axonal damage, and cell birth in the immature rat brain Presented by:Monique Ferguson Bio 475 Del Bigio, M.R, and Y.W. Zhang. 1998. Cell death, axonal damage, and cell birth in the immature rat brain following induction of hydrocephalus. Exp. Neurol. 154:157-169.

2. Derived from Greek words: “hydro”=water “cephalus”=head HydrocephalusWhat is it? • It is a condition in which the primary characteristic is excessive accumulation of fluid in the brain resulting from a blockage in CSF route of flow and reabsorption

3. Cerebrospinal Fluid • The water/fluid is actually CerebroSpinal Fluid (CSF) which is a clear liquid surrounding the brain and spinal cord. • It also fills the ventricles or interior chambers of the brain. • CSF is produced by the choroid plexus http://www.angelfire.com/wa/wafshaf50/CSF.html

4. How to identify Hydrocephalus http://sun.science.wayne.edu/~bio340/StudentPages/Gumma/ When the circulation or absorption of CSF is blocked, or a large amount of fluid is produced, the volume of the brain becomes excessively larger than normal. The picture above shows the flow of CSF in the ventricles of a normal individual in comparison to an individual with Hydrocephalus, displaying enlarged ventricles.

5. Types of Hydrocephalus Congenital Hydrocephalus – present at birth and caused by pregnancy complications or genetic dispositionAcquired Hydrocephalus – develops at the time of birth or later in life • Communicating hydrocephalus • Obstructive hydrocephalus (non-communicating) • Normal Pressure Hydrocephalus or NPH • Dandy-Walker Malformation

6. Types of Hydrocephalus Continued • CommunicatingWhen the flow of CSF is blocked after it exits the ventricles. CSF can still flow between the ventricles which remain open • Obstructive (non-communicating)When the flow of CSF is blocked along one or more of the narrow pathways connecting ventricles • Normal PressureThe CSF is produced in normal amounts in this condition, but it is prevented from being normally re-absorbed because of obstruction to the flow of CSF • Dandy-Walker MalformationRare malformation of the brain that is present at birth and is characterized by an enlarged space at the back of the brain that interferes with the normal flow of cerebrospinal fluid through the openings between the ventricle and other parts of the brain.

7. genetic inheritance developmental disorders tumors blockage of the ventricles by hemorhaging complications of premature birth diseases (meningitis) traumatic head injury What causes Hydrocephalus?

8. Symptoms of Hydrocephalus Symptoms of hydrocephalus vary with: • Age • Disease Progression • Individual differences to CSF

9. vomiting sleepiness seizures irritability sunsetting of the eyes (AKA downward deviation of the eyes) Symptoms in Infants

10. An obvious indication of hydrocephalus in infants is the increase in head circumference. In infants the skull has not fully fused and the accumulation of fluid causes the head to enlarge due to the swelling of the ventricles. http://www.mbi.ufl.edu/~joneslab/hcpix.html

11. Headache Vomiting Nausea Papilledema (swelling of the optic disk which is part of the optic nerve) Slowing or loss of development Lethargy Drowsiness Blurred vision Diplopia (double vision) Sunsetting of the eyes Problems with balance Gait disturbance Urinary incontinence Irritability Changes in personality or cognition including memory loss Symptoms in older children and adults

12. How is Hydrocephalus diagnosed • Clinical neurological evaluation • Cranial imaging techniques -ultrasonography -computed tomography (CT) -magnetic resonance imaging (MRI) -pressure monitoring techniques

13. Current Treatment • Shunt System A shunt system diverts the flow of CSF from a site within the central nervous system to another area of the body where it can be absorbed as part of the circulatory system • Third ventriculostomy

14. A shunt is a flexible but sturdy silastic tube A shunt system consists of the shunt, a catheter, and a valve One end of the shunt is placed within the CNS and the other end of the catheter is placed within the abdominal cavity, heart or lung cavity where the CSF can drain and be absorbed The valve located along the catheter maintains one way flow and regulates the rate of CSF flow Shunt System http://www.ohiohealth.com/healthreference/display.htm?image=/images/image_popup/r7_shunt.jpg&caption=A% 20ventriculoperitoneal%20(VP)%20shunt%20(left)%20diverts%20excess%20CSF%20from%20the%20brain%20into%20the%20space%20in%20the%20abdomen%20that%20surrounds%20the%20digestive%20organs%20(peritoneal%20cavity).%20A%20ventriculoatrial%20(VA)%20shunt%20(right)%20diverts%20excess%20CSF%20from%20the%20brain%20into%20the%20right%20atrium%20of%20the%20heart.&title=Hydrocephalus

15. Third Ventriculostomy • Third ventriculostomy involves entering the brain through the bones at the top of the skull. The neurosurgeon passes an neuroendoscope through the lateral ventricle into the third ventricle and uses a laser to make a hole in its floor. Excess fluid drains through the hole into the subarachnoid space. • The overall success rate of third ventriculostomy is about 65%. When used to treat blockage caused by tumor success rates are slightly higher. In hydrocephalus caused by hemorrhaging or infection, they are slightly lower.

16. Cell death, axonal damage, and cell birth in the immature rat brain following induction of hydrocephalus

17. Rationale • To test the hypothesis that hydrocephalus causes death of brain cells and that hydrocephalus is associated with the generation of new brain cells

18. Materials and MethodsExperimental model • 3 week rat with kaolin induced hydrocephalus • Note: At this age the rat brain is developmentally similar to that of a human infant. • Controls received sham injections with needle insertion only

19. Materials and Methods • Histological Studymicroscopic study of the animal tissues • Axonal Transport Study • Electron Microscope Studies • Immunoblot Studies

20. Diagram of rat brain and cerebrospinal fluid system The CSF is secreted in the ventricles by the choroid plexus. CSF flows through the ventricular system (black arrows) and out into the subarachnoid space via openings between the fourth ventricle and the cisterna magna. http://www.mbi.ufl.edu/~joneslab/ratcsf.htm

21. Results • By 2 weeks hydrocephalic rats weighed less than controls • By 3-4 weeks hydrocephalic rats exhibited unsteady gait, poor grooming and domed skulls due to progressive ventricular enlargement

22. Results continued • Reactive changes and Inflammation • Axonal damage • Cell death • Cell generation

23. Reactive changes and Inflammation

24. Anti-Glial fibrillary acidic protein (anti-GFAP) and Nestin immunoreactivity • A=Immunohistochemical labelling of reactive astrocytes. At 3-4 weeks in hydrophalic rats there were abundant reactive astrocytes • B=Nestin immunoreactivity was studied with the hope that it might aid in the identification of progenitor cells B A

25. The function of oligodendrocytes is to form myelin in brain and spinal cord Reactive change in oligodendroglia was manifested immunohistochemically by increased expression of gelsolin in cell processes Gelsolin is involved with the movement of myelin-forming cell processes. Gelsolin immunohistochemical labeling of oligodendrocytes

26. Microglial reaction by immunohistochemical labeling with OX-42 B A • A=OX-42 amtibody in the white matter adjacent to the enlarged lateral ventricle of a rat that has been hydrocephalic for 4 weeks • B=No labeling in a comparable region for the control rat. • C=Plump activated microglial cell and reactiev astrocytes were identitfied in hydrocephalus C

27. Axonal Damage

28. Axons labeled with anti-neurofilament • A=Shows the axons labeled with anti-neurofilament in hydrocephalic rat • B= At higher magnification the damaged axon profiles are round and varicose • C=the majority of swollen axons were found to be myelinated Axons are specialized for rapid conduction of nerve signals. They are a part of the neuron.

29. Damaged Axons in White Matter of Control and Hydrocephalic Rat Brains • The controls had the least number of damaged axons and the smallest ventricle size • As the weeks progressed the number of damaged axons increased and ventricle size increased also Axons are specialized for rapid conduction of nerve signals. They are a part of the neuron.

30. Retrograde transport of FluoroGold • Retrograde transport of FluoroGold was used to asses the integrity of the axons • The number of retrogradely labeled neurons in the contralateral cortex tended to be greater in hydrocephalic rats than in the matched control rats The labeling index estimated the number of contralateral labeled neurons per mm3 of ipsilateral cortex injected with FluoroGold label

31. Cell Death

32. Cell death in a rat that had been hydrocephalic for 4 weeks • A- shows a degenerating glial cell • B- shows a glial cell engulfing a dense apoptic body* B A

33. Cell death in hydrocephalic rats • There is a significant increase in cell death in hydrocephalic brains 3 to 4 weeks after kaolin injection

34. Cell Generation

35. Ki67 Immunoreactivity and Histone mRNA as Indicators of Cell Activity in control and Hydrocephalic Rat brains • Demonstration of histone mRNAs showed many S phase cells. There were no statistically significant differences between hydrocephalic and control brains. • Labeling of Ki67 demonstrated abundant cycling cells in all regions of the control. One week after kaolin injection, the quantitiy of Ki67 immunoreactive cells increased significantlyNo such change was found in severely hydrocephalic rats 3 weeks after kaolin injection

36. Fluorescence photomicrograph showing • In controls and moderately hydrocephalic rats, 30-37% of transferrin-positive cells coexpressed Ki67 Ki67 Transferrin

37. Summary • Kaolin induced hydrocephalus in neurologically immature rats was associated with progressive damage to axons in the periventricular white matter and gradual death of oligodendroglial cells • Multiple brain responses were observed: astroglial hyperplasia and hypertrophy, oligodendroglial reaction, microglial activation, and increased cell proliferation in the subependymal zone. • Oligodendrial cells did not appear to regenerate.

38. Questions?