Cell bio exam 4

Tyrosine kinase receptors , SARS - CoV2 and spike proteins

TYROSINE KINASE RECEPTORS

EGFR receptors - type of RTK Mutated in many forms of cancer

Deletion of Exon19 occurs in ~10-20% of lung cancers

Makes EGFR receptors more active

SARS-CoV-2

Corona virus

Lipid membrane surrounding virus

spike proteins - That bind host receptor (ACE2)

RNA genome

Nucleocapsid protein protects RNA

Causes the disease COVID-19

SPIKE PROTEINS

binds ACE2

changes shape and fuses w cell membrane

THIS IS THE ENTRY MECHANISM

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Attachment and Entry and processing

ATTACHEMENT

Spike protein Binds to ACE-2 receptor

Angiotensin converting enzyme 2

Found in heart, lungs, and testes most prevalently

This is protein-protein interaction → very specific

Cells must have:

ACE2 (receptor)

AND enzymes like TMPRSS2 or CTSL

More enzymes = easier infection/entry

ENTRY AND PROCESSING

ENDOCYTOSIS

Enters through receptor mediated endocytosis

Low pH in lysosome/ endosome free RNA genome from virus

Viral membrane protein facilitate release of genome into cell

TRANSPORT ALONG MICROTUBULES

Flu viral entry is similar to COVID

Endosome travel along microtubules (dynein) Actin plays a role in endocytosis

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Replication and packeging

VIRAL REPLICATION

Host ribosome translate viral RNA

Replication complex

RNA dependent RNA polymerase Replicated genome

Proteases

Cleave proteins to functional viral units

VIRUS STOPS CELL FROM MAKING ITS OWN PROTEINS

Nps1 - viral protein that shuts down host protein production.

Possible that virus is better at attaching to ribosome so that when Nps1 falls out the virus can take over

PACKEGING

Virus makes droplets that house RNA

Virus escapes through a Nsp3 protein

Nsp3 helps: protect viral RNA from immune detection

RNA is replicated Proteins are translated into the ER and move through the Golgi Vesicles from Golgi envelope viral genome Exocytosis occurs

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Nps1 interacting with ribosomes Pink -Nps1

RNA -blue

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Export and cell death

EXPORT

EXOCYTOSIS Influenza is model for this as well

Particles are transported along the microtubules and spit out in vesicles.

Actin plays a role in exocytosis

HOW DO VIRUSES KILL CELLS

Some virus induce cellular degradation as their escape mechanism

Corona virus can induce apoptosis

Some types Due to co-opting of cellular machinery

Specific protein may induce cellular response

Apoptosis (programmed death → controlled)

Necrosis (cell bursts → messy inflammation)

Overusing cell machinery until it collapses SARS-CoV-2 can trigger apoptosis or stress responses, contributing to tissue damage.

Where to target a drug

Where to target a drug (general)

You can block the virus at multiple steps:

Entry (ACE2 binding)

Drugs here stop the virus from even getting inside:

Block spike protein

Block ACE2 interaction

Block TMPRSS2 (host protease)

Protease activity (cutting viral proteins)

RNA replication (RdRp)

Virus makes one long protein → needs proteases to cut it

nsps (like Nsp3) form replication machinery

RdRp copies RNA

Drugs here:

Paxlovid → protease inhibitor

Molnupiravir → RdRp inhibitor

Assembly/release

Why target the spike protein and mRNA vaccines

TARGET THE SPIKE PROTEIN

Spike is glycosylated

Antibodies have a hard time

Convalescent plasma seems to be effective for acute treatment

mRNA vaccines

Body makes the spike protein but nothing else

Natural glycosylation occurs so self made antibodies are better

All vaccines do the SAME goal ----> show immune system spike

mRNA vaccines – how they work

mRNA of spike protein is injected and protein is made

mRNA is degraded just like all your normal mRNA

Immune system is presented with a foreign protein made by mRNA

Vaccines

Vaccines:

Reduce infection risk

MASSIVELY reduce severe disease/death

Variant names

Alpha/beta – first wave

Delta – second wave

Omicron – 3rd wave

Virus evolves by:

Mutations

Recombination (mixing variants)

Mutations mostly occur in:

RBD (receptor binding domain) → affects ACE2 binding

NTD → affects antibody recognition

Spike has two major parts:

S1 → binds ACE2

S2 → fusion

Variant prevalence

Variant prevalence over time

Early → Alpha/Beta

Mid → Delta takes over

Late → Omicron completely dominates

Variants don’t stop evolving.

You’ll see new lineages (XBB, BA, etc.)

Each has slight mutations → affects:

transmissibility

immune escape

Long COVID = symptoms lasting months after infection

long-term damage/response

The more reinfection that occurs the higher the rate of long covid

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DIFFERENCES IN ION CONCENTRATION CREATE MEMBRANE POTENTIALS

DIFFERENCES IN ION CONCENTRATION CREATE MEMBRANE POTENTIALS

Small amount of ion movement can cause large changes in membrane potential (Vm)

Membrane potential = the voltage (electrical difference) across a cell membrane

ions involved are VERY close (<1nm) to the membrane

Electrochemical gradient is created by protons being more concentrated on one of the membrane

2 Parts (concentration AND electrical gradient)

Both pushing the same directions

Both caused by the movement of one ion

Membrane potential gradients

MEMBRANE POTENTIAL IS BALANCE OF VOLTAGE AND CONCENTRATION GRADIENTS

Membrane potential is measured in reference to the extracellular space

Membrane potentials may not have both forces going the same way

concentration and voltage gradients could be going opposite.

Inside cell:

K⁺ HIGH

Na⁺ LOW

Ca²⁺ VERY LOW

Outside cell:

Na⁺ HIGH

Cl⁻ HIGH

Ca²⁺ HIGH

These gradients power:

action potentials

signaling

GENERATING MEMBRANE POTENTIALS

GENERATING MEMBRANE POTENTIALS

NON-SELECTIVE CHANNEL: + ONLY

Channel opens and ions flow

Keep track of relative charge either side of the membrane(grey) and concentration (red)

Net Change:

Out: +3 charge, 3 ions

In: -3 charge, 3 ions

Final result is a balanced membrane

NON- SELECTIVE CHANNEL: + AND -

Both + and − ions move, so charges cancel out. Even though ions move, there’s still no electrical difference, so no membrane potential is generated.

SELECTIVE CHANNEL: + AND -

Only + ions flow down concentration gradient

Only positive ions move → charges become uneven across membrane

This creates a voltage difference = membrane potential

Final result is a voltage gradient

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Anatomy of neuron resting potential and ion channel

Anatomy of a neuron

Dendrites = receive signals

Cell body = integrates signals

Axon = sends signal (action potential)

Terminal = communicates with next cell

Voltage can be recorded in a neuron

We place an electrode in a neuron and measure membrane potential over time → this gives us graphs like action potentials.

Resting potential – voltage difference when the cell is not transmitting a signal

AP is the voltage change in the cell that is transmitted along the axon

Ion channels have 3 states:

Closed → nothing happens

Open → ions flow → voltage changes

Inactivated → temporarily shut off

Depolarization = more positive inside

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What do neurons do? Synapse

What do neurons do?

Detect and response to stimuli

Carry signal from one place to another

NEURONS COMMUNICATE WITH EACH OTHER THROUGH SYNAPSES

Pre neuron → axon → synapse → post neuron

Electrical signal travels down axon

Gets converted to chemical signal at synapse

Then back to electrical in next neuron

SYNAPSE IS CONNECTION FROM NEURON TO ANOTHER CELL

Vesicles containing neurotransmitter gather at the synapse

Signal transmission across cleft (rest vs activated)

Resting nerve terminal

voltage-gated Ca²⁺ channel CLOSED

Activated nerve terminal

voltage-gated Ca²⁺ channel OPEN

arriving action potential (electrical signal)

fused synaptic vesicle

released neurotransmitter (chemical signal)

Ca²⁺ = trigger for release

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Activating a postsynaptic cell

Neurotransmitter binds receptor

activating neurotransmitter receptor

this

Opens ion channels

This causes ions to flow → Changes membrane potential → New signal generated

This is how one neuron activates the next

DOPAMINE

Dopamine released → binds receptors → signal

Transporter pulls dopamine back in (recycling)

👉This is how drugs like cocaine work (block reuptake)

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Communication between neurons

Recording communication between neurons

Electric stimulation in one neuron recorded in the same neuron

Electric stimulation in one neuron recorded from another

Explanation: Two setups:

Same neuron → measures action potential

Different neuron → measures synaptic transmission

Synaptic plasticity – cellular basis for memory

Incoming signal stays the same

Even if input is the same, output can change.

This is learning:

Stronger response = memory forming

Weaker response = forgetting

Terms:

Plasticity – ability of a neuron to change its response to a signal

LTP – long term potentiation – increased response to the same input signal

LTD – long term depression – decreased response to the same signal

Caveats:

Synaptic plasticity is not the same thing as remembering and forgetting

Memories are likely made of groups of neurons cooperating with each other

The word “RED” is not stored in one neuron

Case study intro

Dr. Marshall Westwood ate a meal of pufferfish.

felt numbness everywhere

Started throwing up

As Dr. Westwood was rushed to the hospital, he had: - trouble breathing; - signs of paralysis in his upper body and arms.

THE PHYSICIANS

• kept his airway open.

• administered drugs to bring his heart back to a normal rhythm.

• put a mixture of charcoal into his stomach to help absorb any chemicals that might still be there

he learned that he had probably been the victim of a pufferfish poisoning.

Resting potential in neurons

REMEMBER All cells have an electrical potential difference across their plasma membrane. - Called a MEMBRANE POTENTIAL

• The cell's inside is negative relative to the outside.

• Sodium (Na+) potassium (K+) and large anions (A-) are important in maintaining the membrane potential.

Diffusion =high to low (Na+ diffuses into cell bc conc. is high outside)

Nerst equation (finally math ) RESTING POTENTIAL OF ONE ION

V = (RT/zF)*ln (Co/Ci) Co/Ci = concentration outside / inside

RESTING POTENTIAL IN NEURONS

Diffusion of K+ (and less Na+) leads to a separation of charges across the membrane, and the resting potential. - Remember: MANY K+ and very few Na+ channels, membrane permeability is 100x for K+ than Na+. - Movement of K+ increases the positive charge outside the membrane relative to the inside.

Thus an ELECTRICAL gradient is formed that can also influence ion flow.

Mainting concentration gradient

RESTING POTENTIAL IS WHEN ION FLOW IS IN BALANCE WITH DRIVING VOLTAGE GRADIENT

How does the cell maintain the K+ driving force (aka concentration) if K+ is always leaking out

ACTIVE TRANSPORT

3 Na+ Out of cell 2 K+ In to cell

10-30 fold difference in concentrations from outside to inside

Relative voltage gradient established is: “+” outside and “-“ inside ATP is hydrolyzed for energy

ACTION POTENTIALS: HOW NEURONS COMMUNICATE Neurons use changes in membrane potential for fast communication- called an ACTION POTENTIAL.

• Neurons have voltage gated ion channels, that open or close in response to changes in membrane potential.

• Membrane potential may change in response to stimuli that open or close those channels

We record the wave of ions moving in an axon at a single point

Action potential

At Resting Potential, the inside of the neuron is negative relative to the outside and is thus polarized. (-70Mv).

Na+/K+ pump and K+leak channels maintain this potential

WHEN A NEURON IS STIMULATED, THE AREA OF THE MEMBRANE AT THE POINT OF STIMULATION BECOMES MORE PERMEABLE TO NA+.

If a cell starts at resting potential (-70mv), and then is stimulated:

The membrane voltage will become >-70mV because Na+ will move INTO the cell.

For an action potential (nerve firing) to occur, the cell membrane potential must reach a threshold value of ... ~ -55mV

• When this threshold is reached, the voltage-gated Na+ channels open, allowing more Na+ to diffuse rapidly into the cell. - DEPOLARIZATION

REESTABLISHING RESTING POTENTIAL AFTER AN AP

AT THE PEAK OF THE ACTION POTENTIAL, NA+ VOLTAGE-GATED CHANNELS CLOSE, AND K+ VOLTAGE-GATED CHANNELS OPEN IN RESPONSE TO POSITIVE MEMBRANE POTENTIAL. TO RETURN THE CELL TO ITS NEGATIVE RESTING POTENTIAL QUICKLY:

K+ will diffuse along its concentration gradient OUT of the cell

Returns back to resting

back to the case study

Tetrodotoxin in fish is a neurotoxin - it affects nerve cells (neurons).

• Specifically, tetrodotoxin blocks voltage-gated sodium ion channels.

This can lead to Paralysis bc now there is no action potential so theres no signal to muscles

Then after recovering he touched a bird

Dr. Westwood got same symptoms but they quickly faded

It was a pitohui

he isolated the toxic compound that he was active in the feathers of the pitohui. It was a homobatrachotoxin

Batrachotoxin and homobatrachotoxin

Batrachotoxin and homobatrachotoxin are both known to act on voltage-sensitive sodium channels in excitable tissues.

THEY Prevent Na+ ion channels from closing so after depolarizing , they cant repolarize.

Na/k channels

Na⁺ channels open when voltage threshhold is met and inactivate at peak.

K+ channels do the opposite

GPCR FAST EFFECTOR RESPONSE

G proteins can effect ion channels to have immediate effect Beta/gamma interacts with K+ channel and allow K+ to flow across membrane Gi G-protein

Heart pacemakers cells have a mechanism that causes action potentials to fire on a regular basis by leaking in + ions slowly.

What happens to regular pace of action potentials when K+ channels are activate by Gi linked GPCR?

K+ leaks out of the cell canceling the movement of Na+, lowering the membrane potential, and slowing the rate of action potential firing because it is harder to reach threshold

DECREASED RATE OF FIRING

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Mice shhh

CHANNELRHODOPSIN

Scientists put a light-activated ion channel into neurons

Shine blue light → channel opens → Na⁺ goes IN

That causes depolarization → neuron fires

ACTIVATING NEURONS ALTERS BEHAVIOR

Parts of Neuron

Neuron parts are specialized:

Each uses different structural proteins

Different areas of the cell have different cytoskeletal elements

Soma = cell body

Nucleus • Mitochondria • Ribosomes • Etc

Cytoskeleton

Actin ,Microtubules, Neurofilaments

Neurofilaments relate to diseases like Alzheimer’s

Axon hillock

Initial segment for the axon

Cytoskeletal elements compress into axon

Free of most organelles

Due to compression of cytoskeleton limiting the space available

Axons

Only 1 axon present in each neuron

Has MTs and neurofilaments

Tau is the spacer for microtubules - determines spacing

Transport of mitochondria and other elements

No ER or Golgi

Contd

Dendrites

Receive information from other neurons

• Synapses

• More widely spaced cytoskeletal elements that allow for diffusion

• MAP2 - •

Relatively fewer neurofilaments • Thousands of connections

Material transport

Material must be transported around the cell - axons make this more difficult

• Small molecules diffuse down the axon

• Peptides must be transported down the axon

Motor proteins

Synthesis occurs in the soma

• Material is transported along Mts

• Anterograde - towards the synapse - kinesin

• Retrograde - away from the synapse - dynein

Signaling for protein synthesis

Signaling for protein synthesis

Activated PKA can be transported to the nucleus •

PKA phosphorylates transcription factor proteins

DOES MEMORY REQUIRE PROTEIN SYNTHESIS

Short-term memory → NO

Long-term memory → YES

Memory happens in stages:

Learn something

Short-term memory (temporary)

Consolidation → long-term memory

Behavioral testing for memory

Train an animal to have a response to a certain set of conditions.

Limitations in model animals

Response repertoire

Size and repeatability of response

Fear conditioning

Strong repeatable responses

Variable strength of memory

Present and measurable with non-verbal behavior in many model systems

Zebrafish - twitching

Mouse - freezing and jumping

Fish experiments

Design a test that would determine if protein synthesis is required for different memory types

Block protein synthesis

Test memory later

Memory works at 1 day → short-term ok

Memory fails at 4 days → long-term needs proteins

Classic conditioning:

Light → shock

Fish learns: light = danger

Test memory after time

If fish moves away when light turns on: → memory exists

Short-term memory:

No protein synthesis

lasts hours-day

Long-term memory:

Requires protein synthesis

requires gene transcription

lasts days+

Autophagy vs proteasome

PROTEIN NEEDS TO BE REMOVED DUE TO DAMAGE OR REGULATION OF ACTIVITY

Proteasome - direct degradation

Short lived proteins misfolded monomers

Autophagy - delivery to lysosome Long lived proteins misfolded oligomers/monomers

Both are tagged with ubiquitin

Autophagy is for

nutrient and energy homeostasis

removal of damaged organelles

removal of aggregate proteins

removal of intracellular pathogens

Macroautophagy

molecules are enveloped by a membrane to form autophagosome

membrane fuses with lysosome

Microautophagy small debris are sequestered into lysosome

Chaperone Mediated Autophagy (CMA) proteins are transported through a channel

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PROCESS OF AUTOPHAGY and Phagophore formation

1. Phagophore or isolation membrane forms from membrane 2. Cargo is brought to autophagosome

3. Autophagosome fuses with endosome (amphisome)

4. Amphisome fuses with lysosome (autolysosome)

PHAGOPHORE FORMATION

All autophagosomes start as an isolation membrane (IM) could come from Golgi, endosomes, ER, mitochondria associate membrane (MAM) interface

Complex mechanisms enable formation of autophagic vesicle

PI3P

VPS34 complex

ATG9 - membrane receptor

LC3 - autophagy receptor

Vps34 is phosphotidylinositol kinase (PI3K)

each version of PIP has a different kinase

Turns PI to PI3P PI3P is sensed by other proteins

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Autophagosome formation and LC3 - II

ATG9 containing vesicles bud from source membrane Vesicles assemble into preautophagosomal structure (PAS) PAS forms isolation membrane

ATG9 senses PI3P to bring new membrane to forming autophagosomes

LC3-II IS A PHOSPHOLIPID TAG FOUND ONLY IN AUTOPHAGOSOMES

LC3-II is lipidated form of LC3-I

lipidation is when PE is added to LC3

LC3-II is a molecule used to “tag” PE

Found only in autophagosome

PE part of LC3-II can be inserted to membranes to allow ‘tag’ to be associated with membrane

Autophagy regulation

Starvation induced autophagy

TOR complex inhibits autophagy

anything that inhibits TOR will activate autophagy and vice versa

insulin and amino acids activate TOR Class III PI3K (kinase) activates autophagy

ATG proteins - autophagy related work with kinase to form protein complexes

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Mitophagy

CARGO IS LOADED INTO PHAGOPHORE AND BECOMES THE ISOLATION MEMBRANE

MITOPHAGY

Damaged mitochondria ubiquinated

Ubiquitin recognizes LC3II (potentially using an adaptor protein P63)

Mitochondria is loaded into the auotphagosome

Pros

Aging - removes damaged organelles

Neurodegeneration - prevents accumulation of aggregate proteins

Infection - this is one of the way that cells get rid of viruses and bacteria that get inside

Cancer - removes damaged organelles

cons

Neurodegeneration - accumulation of autophagic vesicles may be involved in AD

Infection - some bacteria have evolved to subvert the system for their lifestyle

Cancer - can help cancer cells resist cancer drugs and survive with low nutrients

Extracellular matrix

EXTRACELLULAR MATRIX

Supports and anchors cells

Cell communication

Composition:

Mix of proteins and polysaccharides

Secreted from cells via exocytosis

COLLAGEN

Single protein chain

Becomes triple strand

Strands assemble into fiber

Fibers assemble into bundles

PROTEOGLYCANS

GAGs made of repeated sugar elements

Keratin, Chondrotin sulfate, Hyaluronan

Proteins link together GAGs

LAMININ

Fibrous proteins

INTEGRINS

Used to connect cells to extracellular matrix

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CELLULAR CONNECTIONS

CELLULAR CONNECTIONS

tight junction → seals neighboring cells together in an epithelial sheet to prevent leakage of extracellular molecules between them; helps polarize cells

adherens junction → joins an actin bundle in one cell to a similar bundle in a neighboring cell

desmosome → joins the intermediate filaments in one cell to those in a neighbor

gap junction → forms channels that allow small, intracellular, water-soluble molecules, including inorganic ions and metabolites, to pass from cell to cell

hemidesmosome → anchors intermediate filaments in a cell to the basal lamina

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Tissues , Epithelial

TISSUES

TISSUE ARE MADE OF MULTIPLE TYPES OF CELLS

Organs = multiple tissue layers working together

Skin is NOT just one cell type

It has:

epithelium (top layer)

connective tissue (middle)

fat (bottom)

Epithelial types

columnar

cuboidal

stratified

squamous

basal lamina

Epidermis

top = dead protective layer

keratinocytes → structure

melanocytes → pigment

Langerhans → immune

Loose connective tissue = spread out cells + immune cells

immune defense

flexible

dense connective tissue of dermis

Dense connective tissue = tight fibers

strong (tendons, dermis)

less space, fewer cells

QUORUM SENSING

Allow bacteria to sense the density of their population

An auto inducer is produced by the bacteria constitutively

The bacteria have a receptor that binds with the auto inducer and starts a signaling cascade

Extracellular concentration < intracellular concentration

Cells continue to replicate

Replicated cells also begin to produce autoinducer (AI)

Intracellular AI stops moving out of the cell

Higher AI concentration downregulates itself by inhibiting its own transcription factor

WHAT ARE EXTRA CELLULAR VESICLES?

Lipid bi-layer vesicles critical for cellular communication

Released from lots of different kinds of cells - plants, nematodes, mammalian, microbes

Can not replicate

Carry proteins, nucleic acids, metabolites, organelles

Initially thought to be a way the cell got rid of trash

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Extracellular vesicles

WHAT ARE EXTRA CELLULAR VESICLES?

Lipid bi-layer vesicles critical for cellular communication

Released from lots of different kinds of cells - plants, nematodes, mammalian, microbes

Can not replicate

Carry proteins, nucleic acids, metabolites, organelles

Initially thought to be a way the cell got rid of trash

3 types

Microvesicles

Exosomes

Apoptotic bodies

WHAT IS IN AN EV?

Proteins

Nucleic acids

Lipids

HOW ARE EVs RECEIVED BY OTHER CELLS

Phago/pinocytosis

membrane fusion

Scientists still don’t fully understand EVs

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How are they received

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Case study again intro

Jackie got Botox → develops drooping eyelid (ptosis)

Cause: Botox spread to the levator palpebrae superioris muscle

That muscle normally lifts the eyelid

Botox = Botulinum neurotoxin (BoNT)

A neurotoxin that blocks neurotransmitter release by cleaving SNARE proteins.

It:

Enters neurons via receptor-mediated endocytosis (clathrin)

Gets into cytosol

Cleaves SNARE proteins

Blocks synaptic vesicle exocytosis

Prevents acetylcholine (ACh) release

SNARE proteins

Proteins that allow vesicle fusion (synaptobrevin, syntaxin, SNAP-25)

No ACh → no muscle contraction → paralysis

More

Botox enters cells like LDL

Uses receptor-mediated endocytosis

Specifically: clathrin-dependent

Gets into cytosol

Targets SNARE proteins

Blocks exocytosis of ACh

Neuron cannot release acetylcholine-> Muscle does not contract = paralysis

Why are endocytosis and exocytosis active transport?

Because they require ATP (energy)

What is clathrin and its role?

Clathrin = protein that forms coated vesicles

Examples of proteins secreted by exocytosis

hormones (insulin)

neurotransmitters

enzymes

antibodies

Order

Correct order:

SNARE proteins interact

Vesicle fuses with plasma membrane

Acetylcholine is released from vesicle

Acetylcholine binds to the acetylcholine receptor

Receptor changes conformation

Part 2

ACh binds to nicotinic acetylcholine receptors (nAChRs), which are ligand-gated ion channels, on skeletal muscle.

Binding of the ligand acetylcholine to the nAChRs induces these ion channels to open, allowing Na+ ions to rush into the cell

Channel opens → Na⁺ rushes INTO muscle cell

Membrane depolarizes → triggers contraction

This is facilitated diffusion, not active transport

it's a CHANNEL (not a transporter)

Ligand-gated ion channel

Allows Na⁺ diffusion

Muscarinic ACh receptor (GPCR pathway)

Steps:

ACh binds mAChR (GPCR)

Activates G protein (Gi type)

Effects:

↓ cAMP (inhibits adenylate cyclase)

Opens K⁺ channels

Cell becomes more negative (hyperpolarized)

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Generally, after neurotransmitters are released in the neuromuscular junction, many of the neurotransmitters or some of their components are recycled by a reuptake mechanism inherent in the pre-synaptic neuron. Considering that the reuptake mechanism is often a sodium-dependent process that utilizes membrane potential, what type of membrane transport might you expect to be in play here?

secondary active transport (symporter)

Na⁺ moves down gradient

Neurotransmitter moves with it into neuron

LAST PART

lidocaine,

Uncharged lidocaine diffuses into neuron

Becomes protonated (charged) inside

Binds inside Na⁺ channel

Physically blocks Na⁺ entry

Ach cant enter

No depolarization AND NO nerve signal

GENERAL ANESTHETICS

Don’t block one specific protein — they mess with the membrane itself

Mechanism:

Insert into lipid bilayer

Change:

membrane fluidity

membrane pressure

This alters ion channel shape → channels don’t open properly

Reduced neural activity (brain-wide)

WHY LOPIDINE HELPS

Botox paralyzes levator palpebrae (skeletal muscle)

Lopidine activates smooth muscle (tarsus superior)

Mechanism:

α-adrenergic agonist (GPCR)

Causes contraction → lifts eyelid slightly (~2 mm)

questions

Hydrophobic molecule in membrane — mimic anesthetic?

YES (likely)

Why:

General anesthetics work by inserting into membrane

Your molecule:

spans bilayer

disrupts lipid organization → could alter protein function

IV Botox (super important concept)

This would be VERY BAD

Effects:

Widespread paralysis

Respiratory failure (diaphragm)

No neuromuscular signaling

Why lidocaine stays inside cell

It’s about charge

Outside: uncharged → diffuses in

Inside: becomes protonated (charged)

Charged = can’t cross membrane

So it gets trapped inside neuron

α-latrotoxin (black widow toxin)

Observed:

First: muscle spasms

Then: paralysis

Opposite of Botox

Causes massive ACh release

Likely:

increases Ca²⁺ influx OR

forces vesicle fusion (SNARE activation)

Result:

Overstimulation → spasms

Neurotransmitter depletion → paralysis