class 12: naturally occurring cell death

what developmental purposes are served by programmed cell death?

• cell population regulation & numerical matching

• tissue remodeling & strutcural shaping

• elimination of compromised lineages

• circuit refinement

cell population regulation & numerical matching

overproduction of neurons followed by selective elimination adjusts neuronal numbers to match the size of their target tissues and synaptic circuits

tissue remodeling & structural shaping

during early development, selective apoptosis helps shape early neural structures and facilitates tissue remodeling

elimination of compromised lineages

programmed cell death selectively removes damaged progenitor cells, preventing expansion of defective cell lines

circuit refinement

programmed cell death helps match specific neuronal sub-populations, such as adjusting inhibitory interneuron numbers to match excitatory cortical networks

trophic support withdrawal

loss of survival signaling increases activity of proapoptotic BH3-only proteins

regulation by BCL2-family proteins

anti-apoptotic proteins retrain pro-apoptotic family members.

• upon trophic withdrawal, BH3-only proteins promote activation of pro-apoptotic BCL2-family proteins BAX and BAK

mitochondrial permeabilization

activated BAX / BAK induce mitochondrial outer membrane permeabilization, causing release of cytochrome c from mitochondria

apoptosome assembly & initiator caspase activation

cytosolic cytochrome c triggers assembly of APAF1-containing apoptosome complex, which recruits and activates initiator caspase 9

executioner caspase activation

active caspase 9 cleaves and activates executioner caspase (3 & 7)

• they dismantle cell by cleaving structural proteins and activating regulators of DNA fragmentation

why does the TUNNEL assay unambiguously identify apoptosing cells.

identifies dying cells by labeling exposed 3-OH free ends of DNA

• TUNEL alone cannot serve as a diagnostic, it must be combined with aditional markers such as cleaved caspase 3

explain how apoptotic cells are recognized and removed

• microglia in CNS recognize phosphatidylserine as an “eat-me” signal and rapidly engulf dying cells.

• plasma membrane remains intact until engulfment, preventing release of toxic cellulsr contents and limiting inflammation

why can rapid clearance cause a single tissue section to underestimate the total amount of developmental cell death?

phagocytic engulfment and clearance occur rapidly, because fixed tissue section captures only brief, dying cells disappear quickly from view

• consequently, number of apoptotic profiles visible at any single moment represents small fraction of total cumulative cell death occurring across development.

how can apoptosis contribute to neural tube morphogenesis?

apoptosis occurs around neural folds as neural plate bends and detaches from surface ectoderm.

• removing specific cells provides tissue flexibility and aids in physical remodeling for timely neural tube closure.

explain why observing a closure defect after caspase inhibition does not, by itself, reveal the precise cellular mechanism of the defect.

demonstrates that caspase activity is required, but does not pinpoint precise cellular mechanism

non-apoptotic caspase roles

caspases can execute non-lethal, non-apoptotic functions in cellular signaling and structural dynamics during tissue shaping

indirect tissue mechanics

blocking cell death can alter surrounding tissue cell density rather than disrupting cell-fusion machinery

lack of morphogenetic resolution

static observation of closure failure cannot distinguish whether defect arose from altered cell bending or changes in tissue movement.

describe the role of the transient Subplate neuron in Cortical circuitry

represents a transient cell population that functions as a critical organizer in development in cortical circuitry
- pioneers axonal pathways

predict the effects of limb-bud removal of an limb on the survival of associated motor and sensory neurons

decreases survival of ventral horn motor neurons and DRG neurons by eliminating target support and increasing cell death

predict the effects of transplantation of an additional limb on the survival of associated motor and sensory neurons

increases survival of associated motor and sensory neurons by reducing naturally occurring cell death

explain how limb experiments support the neurotrophic hypothesis and what they do not establish about the identity of the trophic factor.

• support: demonstrate post mitotic neuron production is unchanged, survival depends on competing for limiting quantities of survival factors provided

• not established: tissue-level experiments do not reveal molecular identity of specific trophic factor

in a compartmentalized sympathetic-neuron culture, NGF is supplied only to distal axons. explain how this can support the cell body

when NGF is applied to distal axons, it binds TrkA receptors on axon terminals → internalized into signaling endosomes.

• in soma, signal leads phosphorylation of transcription factor CREB in nucleus, activating pro-survival gene transcription

in a compartmentalized sympathetic-neuron culture, NGF is supplied only to distal axons. predict the effect of selectively disrupting retrograde transport.

selectively disrupting dynein or microtubule based transport prevents signaling endosomes from reaching cell body, causing CRED inactivation and somatic apoptosis

what control would help exclude diffusion of NGF to the somatic compartment?

researchers add anti-NGF neutralizing antibodies to central somatic chamber or maintain a fluid pressure gradient

compare the preferred Trk receptors for NGF, BDNF, NT-4/5, and NT-3.

• NGF prefers TrkA

• BDNF prefers TrkB

• NT-4/5 prefers TrkB

• NT-3 prefers TrkC

why might two neuronal populations exposed to the same neurotrophin show different survival responses

they express different combinations or splice isoforms of Trk receptors, possses different intracellular cascades, or at different stages of development

why should p75 not be described simply as a survival receptor?

it lacks intrinsic tyrosine kinase activity

• can either enhance Trk survival signaling or trigger pro-apoptotic death pathways

why can loss of sensory input increase neuronal death in a central circuit

central neurons rely on incoming presynaptic action potentials and synaptic drive for activity-dependent intracellular signaling

• deafferenation removes depolarizing drive and trophic support, increasing neuronal death

why can curare treatment increase motor neuron survival

it blocks nicotinic acetylcholine receptors at muscle target, preventing muscle contraction

relate the location of each manipulation to activity-dependent signaling, axonal branching, and access to trophic support.

in response to silent muscle, motor axon terminals increase axonal branching → gives motor neurons enhanced access to limited muscle-derived trophic factors → rescuing them from natural cell death