Exam 3

Early Development:

Von Baer’s laws: 4 generalizations of vertebrate development

  1. the general features of a large group of animals appears earlier in development than do the specialized features of a smaller group

    ~ All developing vertebrates look very similar after gastrulation and only diversify later

    ~All vertebrate embryos have gill arches, a notochord, a spinal cord, and primitive kidneys

  2. Less general characters develop from the more general, until finally the most specialized appear

    ~early on, all vertebrates have a similar skin. Specializations such as scales, feathers, hair etc develop later

  3. The embryo of a given species, instead of passing through the adult stages of lower (simpler anatomically) animals, depart more and more from them

    ~For example, all embryonic vertebrates have gill arches. These are not the same as adult fish gills. Rather, fish elaborate and develop these structures develop into the eustachian tubes (ear-mouth connection).

  4. Therefore, the early embryo of a higher animal is never like a lower animal but only like its early embryo

    ~Human embryos never pass through a stage where they look like an adult fish or bird. Rather, human embryos, fish embryos, and bird embryos initially share common characteristics and look similar

General stages of development:

  1. fertilization

  2. cleavage

  3. gastrulation

  4. organogenesis

  5. metamorphosis

  6. gametogenesis

The Germ Layers:formed during gastrulation

  • Three distinct regions of the embryo that give rise to differentiated cell types and specified organ systems

The discovery of the primary germ layers

  • 3 individuals established the science of embryology

  • Christian Pander, Karl Ernst von Baer, and Heinrich Rathke

    • pander discovered the germ layers

The Germ layers

  • Three distinct regions of the embryo that give rise to differentiated cell types and specific organ systems

    • ectoderm

      • outer layer= skin, brain, neural crest

    • Mesoderm

      • middle layer = blood, heart, kidney, gonads, bones, muscles, and connective tissue

    • Endoderm

      • inner layer= digestive tube and associated organs including the lungs

How does the blastula become a gastrula which becomes an adult?

  • a complicated series of cell movements and shape changes allow the formation of the axis

Formation of the body axis and organs

  • how do cells know what to become and when do they know?

  • induction and specification

Specification

  • The first sage of commitment of cell or tissue fate during which the cell or tissue is capable of differentiating autonomously (by itself) when placed in an environment that is neutral with respect to the development pathway

    • at the stage of specification, cell commitment is capable of being reversed if placed in another environment that is not neutral

Induction

  • The process by which one cell population influences the development of neighboring cells via interactions at close range

    • this is the environment that can alter specification

After induction and specification comes Differentiation

  • The process by which an unspecialized cell becomes specialized into one of the many cell types that make up the body

Waddington’s Developmental Landscape

  • the end result of these related processes is the production of specific tissues in the right location

Formation of the body axis and organs

  • How do cells know what to become and when do they know

  • Autonomous specification

    • characteristics of most invertebrates

    • differential acquisition of cytoplasmic molecules present in the egg control what the cell becomes

    • invariant cleavage produces the same lineage in each embryo of the species

    • cell type specification precedes cell migration

    • Bottom line: cells cannot change fate!

      • even when some cells are lost

  • conditional specification

    • cell specification depends on enviornment and neighboring cells

    • each cell is pluripotent

      • can be many things

    • Cell fate is restricted based on external cues

    • Found in most vertebrates

    • Bottom line: call fate is malleable and can be altered

      • to a certain point

    • transplanted cells differentiate into tissue that corresponds to new embryonic region

    • cell removal does not cause any defects in animal- other cells can replace them

Cells that will give rise to certain tissues have their fate determined during gastrulation

  • Fate map: Identification of groups of cells in the gastrula that will become a particular tissue in the adult

  • How do we know this?

    • fate mapping studies to label individual cells and track them over time

    • Method 1: fluorescent dye labeling

      • Step 1: inject cells with fluorescent tracking dye (green)

      • Step 2: see where they go!

    • Method 2: Chimeric organisms, i.e. quail chick chimeras

      • Step 1: Transplant cells from quail embryo into chick

      • Step 2: see where they go!

        • how do we identify quail cells in a chick?

          • 1. Diffrences in nuclear DNA condensation

          • 2. Quail-specific antibodies

            • immunohistochemistry!

Chimeric organisms revealed the diversity of tissues derived from neural crest (glia and neurons of the PNS & Enteric nervous system)

  • Neural crest arises from the ectoderm

  • adjacent to developing neural tube

  • Delaminates and migrates away

  • defects in neural crest migration cause cleft palates

what do the neural crest become

  • cell transplantation showed us a lot!

  • transplant neural crest from pigmented chicken embryo into albino

  • Black feathers!

Induction of the nervous system

  • What is induction

    • the process by which one cell population influences the development of neighboring cells via interactions at close range

    • this is the environment that cal, alter cell specification

  • For the nervous system, induction relies on the organizer

Discovery of the Spemann-Mangold Organizer in amphibians

  • disocvered by Hilde Mangold and Hans Spemann

  • Discovered that transplanting cells from the blastopore lip of a blastula to another embryo resulted in the formation of a second axis/nervous system in newts

The organizer: A powerful modulator of embryonic tissue patterning

  • What is it: Piece of tissue in the blastula that can program the cells around it to eventually become the body axis

  • Where is it: forms at the dorsal blastopore lip

    • structure through which cells migrate during gastrulation

  • Why does it form there: convolution of signals from endoderm and ectoderm

    • General set up of xenopus blastula

      • organizer develops from presumptive mesoderm between ectoderm and endoderm

    • First, mesoderm is induced by endoderm

      • removal of middle cells causes top cells to differentiate into mesoderm instead of ectoderm

    • Then, endoderm (known as Nieuwkoop center) induces the organizer

  • How does it work: involutes and becomes mesoderm

    • anterior part involutes first during gastrulation and induces anterior nervous system

    • Posterior part involutes later, forms posterior mesoderm, induces posterior nervous system

    • 1. Initiates movements of gastrulation

    • 2. Self-differentiates

      • into signaling centers

        • Pharyngeal endoderm

        • Prechordal plate(head mesoderm)

        • Notochord

          • rod of mesodermal cells

          • begins developing at 17 days (humans)

          • develops from organizer after involution (gastrulation)

          • Gone by 7-10 weeks

            • example of an embryonic tissue that undergoes programmed cell death

          • necessary for nervous system induction and patterning

    • 3. induces the surrounding mesoderm to form paraxial mesoderm (forms muscle and bone precursors)

    • 4. Induces the ectoderm to form the neural tube

  • The process by which the CNS forms through interactions with underlying mesoderm is known as primary Embryonic induction

  • All vertebrates have an organizer

  • SMO= Spemann-Mangold organizer

Hensen’s node (organizer) and nervous system patterning in chick and huamn

  • hensen’s node at anterior edge of primitive streak

  • cells migrate into primitive streak during gastrulation to form the germ layers

  • transplant node to a new animal

  • induces second neural tube-primitive neurvous system

What factors does the organizer-dirived tissue produce to induce the neural ectoderm?

  • search for factors that induce the nervous system began in the 1930s

    • Hans Holtfreter showed that if you prevent the notochord from developing, you do not get a nervous system

  • how did they know it was a diffusible factor?

    • separated gastrula ectoderm and blastopore lip from newts with filter paper

      • lip contains organizer and notochord procurers

      • no cells could get through, only diffusible, secreted factors

      • nervous system forms

This mysterious mesodermal structure develops during gastrulation and is essential for nervous system development. In humans, it appears at 17 days post-fertilization but is gone by 7-10 weeks of development

This tissue forms at the dorsal blastopore lip and can pattern the entire embryonic axis

What is the nervous system inducing factor?

  • none. Looking for the wrong thing:

    • nervous system is the default state!

  • Inhibition of one critical factor is necessary for ectoderm to become nervous system

    • That factor is: Bone morphogenetic protein (BMP)

      • small protein that activated BMP receptor

      • Leads to phosphorylation of SMAD

      • SMAD goes into nucleus and activates transcription of target genes

How does BMP impact neural induction?

  • bone morphogenetic protein (BMP) is secreted by the mesoderm and induces ectoderm to become epidermis

  • BMP inhibitors from notochord and other organizer derived tissues prevent ectoderm from becoming epidermis- becomes neuroectoderm by default

    • 3 primary BMP inhibitors: Noggin, Chordin, and Follistain

Regional specificity of nervous system patterning

  • Transplanting of different regions of gastrula to host leads to different structures

    • Done by Hilde Mangold’s husband

  • Signaling gradients

Wnt signaling participates in nervous system patterning

  • What is Wnt signaling?

    • secreted small peptide

    • Bindes to Wnt receptors

    • Allows activation of transcription by B-catenin

    • If Wnt signaling is “off” B-Catenin is degraded

How does Wnt impact neural induction

  • Wnt inhibitors are:

    • Cerebrus, Dickkopf, Frzb, IGF

  • Wnt signaling is active in the posterior region

  • BMP is inhibited everywhere

Wnt and BMP inhibitors induce the head nervous system

  • Anterior region of the head is above the pharyngeal endoderm and head mesoderm

    • secrets inhibitors of Wnt

  • Inhibitors of Wnt are critical for induction of Anterior (cranial) nervous system

  • Wnt inhibitors are only in the anterior region

    • Wnt signaling is active in the posterior region

    • BMP is still inhibited notochord secrets BMP inhibitors

Neurulation: the process of transitioning the neural plate to the neural tube

Formation of the Neural Tube

  • Endoderm- viscera (internal organs)

  • Mesoderm- muscle and bone

  • Ectoderm- nervous system, neural crest, and skin

  • Primary neurulation vs secondary neurulation

  • Primary: cells surrounding the neural plate direct the plate cells to proliferate, invaginate, and pinch off from the surface

    • forms a hollow tube

  • Secondary: neural tube arises from cells that coalesce into a solid cord that subsequently hollows

    • Eventually forms a hollow tube

    • only occurs caudal to the sacral vertebrae in mammals

Primary Neurulation

steps:

  1. After neural plate formation, edges thicken and move up to form neural folds

  2. Neural groove appears in center of plate

  3. Neural folds migrate towards them embryonic midline

  4. Neural folds fuse creating a hollow tube

Formation of the Neural Tube- the importance of folic acid

  • Neurulation happens within the first few weeks of pregnancy

    • oftentimes before pregnancy is known

  • 1 in 500 births has neurulation defects due to nutritional deficits

  • Estimates that 90% of the birth defects are due to lack of folic acid

    • exact reason for folic acid importance is unknown

Three Primary Brain Vesicles

  • First steps of neural differentiation are the swelling of the anterior neural tube leading to the formation of the primary vesicles

  • entire brain develops from these primary vesicles

Differentiation of the Forebrain

  • Next steps: Vesicles sprout off the sides of the prosencephalon

    • 2 telencephalic vesicles, 2 optic vesicles

    • residual central structure that remains in the diencephalon

    • Retina develops from the optic vesicle

      • part of forebrain, not PNS

Formation of the retina from the optic vesicle

  • optic vesicles grow and invaginate

  • form the optic stalks and the optic cups

  • these become the optic nerves and the retinas in the adult

  • retinas and optic nerves are derived from the nerual tube

Differentiation of the Telencephalon and Diencephalon

  • Telencephalon:

    • Differentiates into the cerebral hemispheres, olfactory bulbs, basal telencephalon

    • Grows lateral and posterior to cover diencephalon

  • Diencephalon

    • gives rise to the thalamus, and hypothalamus

Differentiation of the Midbrain (mesencephalon)

  • small changes relative to forebrain

  • dorsal surface becomes the tectum

    • latin for roof

  • Floor becomes the tegmentum

  • center remains open as the cerebral aqueduct-connected to ventricular system

  • While simple, contains critical structures

    • tectum - superior and inferior colliculus

      • critical relays for sensory information on route to thalamus

      • superior - recives input from the eye

        • also called the optic tectum

      • inferior - receives input from the ear

    • Tegmentum

      • substantia nigra and red nucleus

        • substantia nigra degenerates in Parkinson’s disease

        • red nucleus - limb movement

        • critical for infant crawling

Differentiation of the Rostral Hindbrian (Rhombencephalon

  • Differentiates into 3 parts

    • pons

    • Medulla oblongata

    • cerebellum

  • Cerebellum and pons develop from the rostral hindbrain

    • metencephalon

  • Medulla develops from the caudal hindbrain

    • Myelencephalon

  • 4th ventricle sits between the dorsal and ventral hindbrain derivatives

Stages of hindbrain development

  • initially, the rostral hindbrain is a simple tube

  • Dorsal-lateral wall of the tube grows dorsally and medially

    • thease are called the rhombic lips

  • this then expands into the cerebellum

  • the ventral wall (below the 4th ventrical) becomes the pons

Differentiation of the Caudal Hindbrain

  • Medulla oblongata develops from the caudal hindbrain

  • ventral and lateral walls swell leaving only a thin layer of non-neuronal cells

  • Medullary pyramids are present on the lateral edges of the ventral medulla

    • thease are white matter tracts

The medulla oblongata: critical for autonomic functions

  • Heart rate

  • Respiration

  • Blood pressure

  • Reflexes

    • vomiting

    • coughing

    • sneezing

    • swallowing

Hindbrain structure-function relationships

  • Cerebellum: movement control

  • Pons: switchboard connecting cerebral cortex to cerebellum

    • 90% of all axons passing through the midbrain synapse on a nucleus in the pins

    • critical relay to the cerebellum to coordinate movement

  • Medulla: carries axons of the corticospinal tract

    • dirrect connection from brain to spinal cord

    • where the medulla joins the spinal cord, thease tracts cross

      • decussation of the pyramids: crossing of axons from one side to the other

Pyramidal Decussation

  • Pyramidal or corticospinal tract- descending axons that pass throuh the midbrain/pons into the spinal cord

    • pass through the medullary pyramidsd- hense pyramidal tract

  • Pyramidal decussation

    • where the axons in the CST cross from ipsilateral to contrelateral

    • occurs near where medulla (hindbrain) joins the spinal cord

  • Explains contralateral control of movement

    • left hemisphere controls right side of body

Lobes of the Human Cerebrum

  • Functions of cortical regions (oversimplification)

  • Frontal lobe

    • reasoning, impulse control, integration

    • also contains motor and premotor cortex for higher order motor control/voluntary movement

  • Parietal lobe

    • processing somatosensory information

  • opccipital lobe

    • vision/visual processing

  • Temporal lobe

    • language/ auditory processing, memory, visual perception

Cerebral Cortex organization

  • common features of cerebral cortex in vertebrates

    • cell bodies of cortical neurons are always arranged in layers or sheets that lie parallel to the surface of the brain

    • most superficial layer (layer 1) has no neurons

      • made of neuronal processes primarily

      • Beneath the pia mater (meninges)

    • At least 1 cell layer has pyramidal cells that have large apical dendrites that extends up to layer 1

  • These characteristics distinguished cortex from other brain regions, e.g. thalamus

General stages of neurogenesis

  • Birth

  • Migration

  • Differentiation

Neurogenesis: the birth of new neurons

  • Occurs at the ventricular surface of the developing brain

    • During early develop this region is very thin

      • two zones:

        • Ventricular zone-contracting ventricular

        • Marginal zone (dorsal)

  • Neural progenitors are the ventricular zone

    • progenitor cell for all neurons and astrocytes of CNS

    • known as radial glial cells

Radical glial cells: not just a scaffold

  • Radical glial cells were known to exist for centuries

  • consistent presence of the cell bodies near the ventricular zone/surface and long projections extending to the superficial surface of the developing neural tissue

Steps of neurogenesis

  1. Radical glial cell extends process to reach towards the pia (connective tissue and blood vessels) at the surface of the brain

  2. Interkinetic nuclear migration-DNA is replicated

  3. Radical glial cell retracts its apically extending arm

  4. cell division

  5. Migration of neuroblast (also called neural precursor) from ventricular surface

Interkinetic nuclear migration

  • process by which the nucleus of a radial glial cell migrates away and back towards the ventricular zone

  • necessary for DNA replication and ultimate cell division

Types of cell division

  • cell division can be symmetrical or asymmetrical

  • symmetrical division

    • happens early in development

    • expands progenitor population

    • during symmetric divisions no neuroblasts (neural precursors) are formed

  • in humans , majority of neocortical neurons born between 5 weeks and 5 months of gestation

  • Peak neurogenesis rate: 250,000 new neurons per minute

Neurogenesis: the birth of new neurons

  • schematic of the development of the primate embryonic neocortex

    • VZ-ventricular zone

    • iSVZ- inner sub-ventricular zone

    • oSVZ- outer sub-ventricular zone

    • IZ- intermediate Zone

    • CP- cortical plate

    • MZ- marginal zone

  • Sub-ventricular zone

    • specific to primates

    • second layer of proliferative progenitors

    • gives rise to upper cortical layers

Neurogenesis: what cells maintain the ability to divide?

  • Daughter cells from an asymmetric division

    • 1 is a progenitor

    • 1 is a neuroblast and will never divide again

  • some progenitor cells are maintained into adulthood

  • vast majority of neurons you are born with are what you have to work with your whole life

What determines if a cell will be a radial glial cell or neuroblast?

  • symmetrical or asymmetrical division

  • compartmentalization of gene products

  • Transcriptional programs

Review of gene expression products

  • Brief refresh

    • DNA is transcribed into RNA in the nucleus

    • RNA is spliced and exported into the cytoplasm

    • RNA is translated into protein via ribosomes

    • Ribosomes are seated on the endoplasmic reticulum

      • rough ER

    • Protein is packaged and actively moved to correct location

Compartmentalization of gene products

  • During division, asymmetric compartmentalization of specific proteins and RNAs leads to fate determination

Transcriptional programs regulate stem cell fate

Once a neuroblast is born, how does it get where it needs to go?

  • Cell migration in the developing cortex

  • Two primary types of migration

    • cortical migration from ventricular zone of dorsal telencephalon to cortex

      • gives a rise to inhibitory interneurons and oligodendrocytes

Cortical migration

  • Migration of neuroblasts (neural precursors) occurs along radial glial fibers

  • During migration, cells have a distinct morphology

    • leading process

    • Nucleus

    • Trailing process

  • Nucleus is pulled into the leading process as the neuron migrates up the radial glial fiber into cortex

Cortex assembles “inside out”

  • first cells to migrate from the ventricular zone are the subplate cells

    • what is the subplate?

      • transient structure

      • contains a variety of neurons

      • More are glutamatergic

      • responsib;e for early motor behavior

      • activity onset 9-10 weeks post-conception in human

      • Thickest 28-34 weeks post-conception

      • most active fetal brain area

    • what does the subplate do?

      • unclear

      • evidence points to a role in defining cortical regions for sensory processing

    • subplate is gone by 3 months post-birth

Cortex assembles “inside out”

  • Next cells to migrate make up layer VI

  • Then layers V, IV, III, II, I

    • inside out assembly

      • inner layers are set up first and cells migrate past them to form the next layer

  • Mutations can disrupt this assembly

    • Reelin mutant

What is reelin and what does it do?

  • Reelin is an extracellular matrix glycoprotein (sugar-bound protein)

    • protein bound by oligosaccharides

  • secreted by Cajal- Retzius neurons in the marginal zone

  • Binds to very low density lipoprotein receptor (VIdIr)

    • VIdIr- reenlin binding is a stop signal for migration

      • stops cells during initial set up of cortical layers

How was reelin discovered?

  • Spontaneous mutation in a mouse line

    • Named reelin because of its abnormal gate

    • Histopathological analysis revealed deficits in cortical laminar organization

  • abnormal reelin expression linked to a variety of psychiatric disorders: Schizophrenia, autism, Bipolar disorder, Alzheimer’s

Once a neuroblast is born, how does it get where it needs to go?

  • Cell migration in the developing cortex

  • two primary types of migration

    • cortical migration from ventricular zone of dorsal telencephalon to cortex

      • gives rise to pyramidal neurons and astrocytes

    • Tangential migration from ventricular zone of the ventral telencephalon to cortex

      • gives rise to inhibitory interneurons and oligodendrocytes

Tangential migration

  • gives rise inhibitory interneurons and oligodendrocytes

  • Does not use radial glia

    • neurons maintain a stellate shape and are highly individually motile

  • Migrate from the ganglionic emineces

    • medial ganglion eminence

      • produces GABAergic interneurons that migrate to cortex

    • lateral ganglion eminence

      • produces GABAergic interneurons that migrate in the rostral migratory stream to the olfactory bulb

    • Caudal ganglionic eminence

      • also produced inhibitory interneurons that migrate to cortex

Neuronal differentiation

  • occurs immediately following neural precursor (neuroblast) migration

  • Layer IV neurons differentiates before layer III neurons migrate through

  • Stepwise

    • neurite outgrowth

    • Axon and dendrite specification

    • Target selection and stabilization

    • Synapse formation

  • Differentiation is regulated by intracellular and extracellular signals

  • Cultured hippocampal neuron after 8 days in vitro (DIV)

    • Expressing Green Fluorescent Protein (GFP)

The discovery of adult neurogenesis

  • originally thought no new neurons were bon after adulthood

  • 1965 Altman and Das provided evidence that adult neurogenesis occurs in mammals

    • used radiolabeled thymidine (T)

    • Incorporates into DNA during replication

    • Slice brains and expose film

    • Radiation will work like light to create “exposure” of the film

The discovery of adult neurogenesis

  • 1983: Goldman and Nottebohm showed adult neurogenesis in canaries

  • Again with radiolabeled thymidine