Making connections in the brain
The human brain has a multitude of functions. As well as organising our thoughts, feelings and actions, it also controls bodily processes like fertility. At puberty, the brain is ultimately responsible for switching fertility on. The decision-making process is very complex – it depends on many factors such as the age of the individual, whether they are well nourished and whether are they going through a stressful time.
Making neurons glow
Dr Rebecca Campbell (University of Otago) discusses the importance of fluorescent molecules in confocal laser scanning fluorescence microscopy (‘confocal microscopy’) of cells. She explains how green fluorescent protein (GFP) from jellyfish can be used to make specific neurons glow green.
Point of interest: Look out for the fluorescent jellyfish in this clip!
Jargon alert: Confocal microscopy is a specialised form of optical microscopy that makes it possible to take pictures of many thin slices in a sample without actually slicing the sample up. The individual slices can then be built up into a three-dimensional model.
Jargon alert: Green fluorescent protein (GFP) is the most widely used fluorescent protein in microscopy. When viewed using a confocal microscope, cells containing GFP glow bright green in the parts of the cell where GFP is found. Scientists use GFP and similar fluorescent proteins in many different ways – to label specific types of cell (as Rebecca does), to label individual structures within cells and to look at changes in living cells over time.
Dr Rebecca Campbell from the University of Otago is fascinated by how fertility is switched on and off. She studies a network of cells in the brain that control fertility. Rebecca is especially interested in how these cells, called gonadotropin-releasing hormone (or GnRH) neurons, ‘talk’ to each other and to other cells. She uses microscopes to look closely at the shape of GnRH neurons within mouse brains and to trace the connections between them. Rebecca hopes that her work might eventually lead to treatments for infertility.
Neurons under the microscope
Microscopy is crucial for Rebecca’s work because it’s the best way to understand the shape of the cells she studies. Neurons have a very complex shape, with many long skinny branches (called axons and dendrites). They also form a complicated network through their interactions with each other, and microscopy is the best way of studying these networks.
Confocal microscopy of neurons
University of Otago neuroendocrinologist Rebecca Campbell explains why she uses the confocal microscope to look at cell components.
Rebecca’s microscope of choice is the confocal laser scanning fluorescence microscope. This specialised light microscope uses light from lasers to highlight specific parts of a sample – in Rebecca’s case, the branches of GnRH neurons. Only fluorescent molecules show up under the laser light, which makes them easy to find. Confocal microscopy also allows Rebecca to see an entire cell, even a very long skinny one like a neuron, under the microscope.
Finding GnRH neurons in the mouse brain
One of the challenges of studying GnRH neurons is finding them! The mouse brain contains 75 million interconnected neurons, and only about 1,000 of those are GnRH neurons. To make it possible to find individual GnRH neurons under the microscope, Rebecca uses a family of transgenic mice made by members of her research group. These mice are normal except for one thing: their GnRH neurons contain a protein called green fluorescent protein (GFP), which glows green when illuminated with a particular wavelength of light.
When slices of brain from the transgenic mice are illuminated with lasers under the confocal laser scanning fluorescence microscope, the GFP in the GnRH neurons glows bright green. No other neurons are visible, so the GnRH neurons are easy to find.
How GnRH neurons communicate: solving the puzzle
Rebecca’s work has solved a puzzle about how GnRH neurons work together to control fertility. GnRH neurons are scattered throughout the brain and appear disconnected from each other, so scientists didn’t understand how they could form a network. Rebecca and her team changed all this by showing that GnRH neurons actually have unusually long dendrites – more than 2 mm, so the cells are 100 times longer than they are wide! The long dendrites mean they have lots of connections with each other, despite appearing to be distant from each other.
The shape of GnRH neurons
In the transgenic mice used by Dr Rebecca Campbell’s team, GnRH neurons in the brain give off a green fluorescent glow. The yellow/red-coloured neuron has also been filled with neurobiotin (which glows red) to show how long its dendrites are.
How did Rebecca’s team see the true length of GnRH dendrites for the first time? In their brain slices from their transgenic mice, they found GnRH neurons (glowing green) and filled them with a small protein called neurobiotin. This is a tricky procedure that requires a steady hand! After the cells were filled, they could see where the neurobiotin was under the microscope because it glowed red under laser light.
Because neurobiotin is small, it spread all around the inside of the cells. It even made it all the way to the end of the neuron’s very long dendrites. This meant that the whole length of the dendrites could be seen glowing red under the microscope. Before Rebecca’s experiments, scientists had only looked at the distribution of GnRH peptide, which doesn’t get into all the cell’s nooks and crannies. This is why the cell shape appeared to be less complex than it was.
Looking at neurons in 3D
Brain cells in 3D
Dr Rebecca Campbell (University of Otago) explains how she builds three-dimensional computer models of whole neurons (brain cells). Rebecca describes why images from confocal laser scanning fluorescence microscopy (confocal microscopy) are a good starting point for building the models. She talks about how models help her to understand the shape of neurons and how they connect with each other.
Jargon alert: Confocal microscopy is a specialised form of optical microscopy that makes it possible to take pictures of many thin slices in a sample without actually slicing the sample up. The individual slices can then be built up into a three-dimensional model.
One of the goals of Rebecca’s research is to understand what the network of GnRH neurons looks like. This would be an almost impossible task without some way of visualising the network in 3D. Like most microscopes, the confocal microscope only takes 2D images, but these images can be built up into a 3D computer model of a GnRH neuron. Using these 3D models, Rebecca has shown that dendrites from several GnRH neurons come together in intertwined bundles. She and her team are now exploring whether these bundles affect how GnRH neurons control fertility.
Nature of science
Scientific knowledge can change over time when new information comes to light. For instance, when an experiment is done in a new and different way, it can provide evidence that challenges previous conclusions. Rebecca’s team used a new technique to find and study GnRH neurons under the microscope. They changed scientific thinking by showing that the neuron shape was more complex that had previously been thought.
Related resources
Use this interactive to find out more about how transgenic animals are made.
In the Modelling animal cells in 3D activity, students make 3D models of specialised animal cells, imitating what can be seen under high-resolution microscopes.
Useful links
Download this PDF poster Te Reo Māori i Te Ao Rangahau Roro – The Māori Language in the World of Brain Research from Brain Research New Zealand.
An extensive and ‘easy to understand’ glossary of neuroscience and brain terminology is available online and as PDF to download at the Dana Foundation. The Dana Foundation coordinates the global Brain Awareness Week campaign.
