The ''zona limitans intrathalamica'' (ZLI) is a lineage-restriction compartment and primary developmental boundary in the

vertebrate Vertebrates () comprise all animal taxa within the subphylum Vertebrata () (chordates with backbones), including all mammals, birds, reptiles, amphibians, and fish. Vertebrates represent the overwhelming majority of the phylum Chordata, with c ...

forebrain In the anatomy of the brain of vertebrates, the forebrain or prosencephalon is the rostral (forward-most) portion of the brain. The forebrain (prosencephalon), the midbrain (mesencephalon), and hindbrain (rhombencephalon) are the three primary ...

(which is analogous to the human

cerebrum The cerebrum, telencephalon or endbrain is the largest part of the brain containing the cerebral cortex (of the two cerebral hemispheres), as well as several subcortical structures, including the hippocampus, basal ganglia, and olfactory bulb ...

) that serves as a signaling center and a restrictive border between the

thalamus The thalamus (from Greek θάλαμος, "chamber") is a large mass of gray matter located in the dorsal part of the diencephalon (a division of the forebrain). Nerve fibers project out of the thalamus to the cerebral cortex in all directions, ...

(also known as the dorsal thalamus) and the

prethalamus The subthalamus or prethalamus is a part of the diencephalon. Its most prominent structure is the subthalamic nucleus. The subthalamus connects to the globus pallidus, a basal nucleus of the telencephalon. Structure The subthalamus is located ...

(ventral thalamus). Sonic hedgehog (shh) signaling from the ZLI is crucial in the development of the '' diencephalon'', which develops into the

, the pretectum, and the anterior tegmental structures. The ZLI together with the prethalamus and thalamus make up the mid-diencephalic territory (MDT). Labeled cichlid forebrain diagram with shh expression

Labeled cichlid forebrain diagram with shh expression

Discovery

Cell lineage restriction boundaries, across which replicating cells cannot migrate, were first discovered in

invertebrate Invertebrates are a paraphyletic group of animals that neither possess nor develop a vertebral column (commonly known as a ''backbone'' or ''spine''), derived from the notochord. This is a grouping including all animals apart from the chordate ...

s, where the expression of various

Hox gene Hox genes, a subset of homeobox genes, are a group of related genes that specify regions of the body plan of an embryo along the head-tail axis of animals. Hox proteins encode and specify the characteristics of 'position', ensuring that the cor ...

s in each compartment confer the differentiation of observable segments in the adult body of

Drosophila melanogaster ''Drosophila melanogaster'' is a species of fly (the taxonomic order Diptera) in the family Drosophilidae. The species is often referred to as the fruit fly or lesser fruit fly, or less commonly the " vinegar fly" or "pomace fly". Starting with ...

. Analogous structures were discovered in the developing

brain A brain is an organ that serves as the center of the nervous system in all vertebrate and most invertebrate animals. It is located in the head, usually close to the sensory organs for senses such as vision. It is the most complex organ in a ve ...

Rhombomere In the vertebrate embryo, a rhombomere is a transiently divided segment of the developing neural tube, within the hindbrain region (a neuromere) in the area that will eventually become the rhombencephalon. The rhombomeres appear as a series of ...

s, which extend down the embryo from the

hindbrain The hindbrain or rhombencephalon or lower brain is a developmental categorization of portions of the central nervous system in vertebrates. It includes the medulla, pons, and cerebellum. Together they support vital bodily processes. Metencephal ...

, contain clear boundaries and each express various Hox genes that are necessary for differentiation of structures within the body. More anterior regions of the

were examined in search of other cell lineage restriction boundaries, and multiple potential boundaries continue to be studied (see Developmental Boundaries). Hoxgenesoffruitfly

The importance of these compartments as local signaling centers, areas which chemically influence surrounding tissue, was elucidated by first observing differential

expression Expression may refer to: Linguistics * Expression (linguistics), a word, phrase, or sentence * Fixed expression, a form of words with a specific meaning * Idiom, a type of fixed expression * Metaphorical expression, a particular word, phrase, o ...

of Hox genes in various compartments and second by observing mutant

D. melanogaster ''Drosophila melanogaster'' is a species of fly (the taxonomic order Diptera) in the family Drosophilidae. The species is often referred to as the fruit fly or lesser fruit fly, or less commonly the " vinegar fly" or "pomace fly". Starting with ...

and corresponding

phenotypic In genetics, the phenotype () is the set of observable characteristics or traits of an organism. The term covers the organism's morphology or physical form and structure, its developmental processes, its biochemical and physiological proper ...

(physical) changes. The ZLI was first discovered in the chick using explant and lineage-labeling experiments. In the explant experiments, cells in the region that will become the ZLI, the

, and the

in the chick were removed and placed into separate

cultures Culture () is an umbrella term which encompasses the social behavior, institutions, and norms found in human societies, as well as the knowledge, beliefs, arts, laws, customs, capabilities, and habits of the individuals in these groups.Tylo ...

; the cells continued to grow and retained their identity (the ZLI began

transcribing Transcription in the linguistic sense is the systematic representation of spoken language in written form. The source can either be utterances (''speech'' or ''sign language'') or preexisting text in another writing system. Transcription shoul ...

'' shh'', whereas the

and

did not). Necessity of the ZLI and its corresponding '' shh'' expression were evidenced by lack of thalamic and prethalamic

genetic markers A genetic marker is a gene or DNA sequence with a known location on a chromosome that can be used to identify individuals or species. It can be described as a variation (which may arise due to mutation or alteration in the genomic loci) that can ...

in culture (see Signaling). These experiments confirmed the ZLI as a signaling center. In lineage-labeling experiments, cells were genetically marked, so that each time a labeled cell replicated, its progeny were marked as well. Cells that were marked in the developing ZLI and their progeny remained restricted to the zone. These experiments demonstrated the ZLI as a cell-lineage restriction boundary. Not only a boundary, the ZLI is also a

compartment Compartment may refer to: Biology * Compartment (anatomy), a space of connective tissue between muscles * Compartment (chemistry), in which different parts of the same protein serves different functions * Compartment (development), fields of cells ...

with separate cell lineage restriction boundaries both anterior and posterior of a section of '' shh'' expression. The importance of the ZLI was once more confirmed by

ectopic expression Ectopic is a word used with a prefix, ecto, meaning “out of place.” Ectopic expression is an abnormal gene expression in a cell type, tissue type, or developmental stage in which the gene is not usually expressed. The term ectopic expression is ...

of '' shh'' in other regions of the

, known as the

prosencephalon In the anatomy of the brain of vertebrates, the forebrain or prosencephalon is the rostral (forward-most) portion of the brain. The forebrain (prosencephalon), the midbrain (mesencephalon), and hindbrain (rhombencephalon) are the three primary ...

during development (both the

telencephalon The cerebrum, telencephalon or endbrain is the largest part of the brain containing the cerebral cortex (of the two cerebral hemispheres), as well as several subcortical structures, including the hippocampus, basal ganglia, and olfactory bulb. In ...

and diencephalon), inducing a ZLI-like region that induces thalamic fate.

Developmental boundaries

During

development Development or developing may refer to: Arts *Development hell, when a project is stuck in development *Filmmaking, development phase, including finance and budgeting *Development (music), the process thematic material is reshaped * Photograph ...

in both

s and

s, cell lineage restriction boundaries and signaling centers are formed in order to ensure proper differentiation of the body. Chemical signals, like '' shh'' from the ZLI, are often released from these boundaries and compartments in a

concentration In chemistry, concentration is the abundance of a constituent divided by the total volume of a mixture. Several types of mathematical description can be distinguished: '' mass concentration'', ''molar concentration'', '' number concentration'', ...

gradient (the chemicals are in much higher concentrations closer to the source) and confer identity to flanking regions. Other genes differentially expressed in these flanking regions aid in ensuring proper differentiation (see Signaling). 4 week embryo brain

Many developmental boundaries have been studied: within the forebrain alone, the confirmed cell lineage restriction boundaries are the pallial-subpallial boundary (PSB) dividing the dorsal and ventral telencephalon, the diencephalon-midbrain boundary (DMB) posterior to the ZLI, and the ZLI. The ZLI, like each

rhombomere In the vertebrate embryo, a rhombomere is a transiently divided segment of the developing neural tube, within the hindbrain region (a neuromere) in the area that will eventually become the rhombencephalon. The rhombomeres appear as a series of ...

, serves as an independent

that confers the identity of diencephalon in anterior and posterior regions. Other developmental boundaries serve as cell-lineage restriction boundaries but not signaling centers, while others are signaling centers to and from which cells can migrate. Despite discoveries of cell lineage restriction boundaries and compartments in the brain, many of the regions studied have been disproven as segmental boundaries. These areas have potential as signaling centers, which have influence over the development of neighboring tissues. These boundaries have great influence over other regions of the brain: the placement of the ZLI not only affects the size of adjacent regions but also the size of the

. A posterior shift in the ZLI allows more cells to be allocated to the

. The same is true for other developmental boundaries in the brain and throughout the body: shifts in boundaries responsible for allocating a certain amount of tissue to a certain function result in drastic changes in the adult structure. These boundaries are of crucial importance for proper differentiation.

Formation

Initial axis patterning

After gastrulation, the embryo is completely undifferentiated and requires many different cues to initiate proper differentiation of the body. The top (roof plate, on the dorsal side of the embryo) and bottom (floor plate, on the ventral side) play a crucial role in these first steps: each acts through global signaling (signaling throughout the entire embryo) for

dorsoventral Standard anatomical terms of location are used to unambiguously describe the anatomy of animals, including humans. The terms, typically derived from Latin or Greek roots, describe something in its standard anatomical position. This position prov ...

neural patterning. After completion of the

of the

axis, more local signaling occurs in the developing brain: developmental boundaries such as the midbrain-hindbrain boundary (MHB),

s, and the ZLI aid in

anteroposterior Standard anatomical terms of location are used to unambiguously describe the anatomy of animals, including humans. The terms, typically derived from Latin or Greek roots, describe something in its standard anatomical position. This position prov ...

organization.

Emergence of ''Shh'' expression

Shortly after the beginning of

patterning, Shh is expressed along the basal plate (the bottom) of the embryo, which functions in " ventralization of the

neural tube In the developing chordate (including vertebrates), the neural tube is the embryonic precursor to the central nervous system, which is made up of the brain and spinal cord. The neural groove gradually deepens as the neural fold become elevated, ...

, promotion of growth and proliferation, and formation of the

hypothalamus The hypothalamus () is a part of the brain that contains a number of small nuclei with a variety of functions. One of the most important functions is to link the nervous system to the endocrine system via the pituitary gland. The hypothalamu ...

". As the embryo continues to

develop Develop or DEVELOP may refer to: * ''Develop'' (magazine), a trade publication for the video game industry * ''Develop'' (Apple magazine), a technical magazine formerly published by Apple Computer * Develop (chess), moving a piece from its origina ...

, ''Shh'' expression characteristic of the ZLI extends

dorsally Standard anatomical terms of location are used to unambiguously describe the anatomy of animals, including humans. The terms, typically derived from Latin or Greek roots, describe something in its standard anatomical position. This position prov ...

to form a wedge that eventually narrows to a strip at approximately 22

somite The somites (outdated term: primitive segments) are a set of bilaterally paired blocks of paraxial mesoderm that form in the embryonic stage of somitogenesis, along the head-to-tail axis in segmented animals. In vertebrates, somites subdivide ...

s (the number of developed

s) or less than one day in

zebrafish The zebrafish (''Danio rerio'') is a freshwater fish belonging to the minnow family (Cyprinidae) of the order Cypriniformes. Native to South Asia, it is a popular aquarium fish, frequently sold under the trade name zebra danio (and thus often ca ...

. Although ''Shh'' expression extends

from the basal plate, the ZLI is capable of forming even without the basal plate or mesodermal tissue. '' Shh'' cooperates with ''dlx2'' and ''fezl'' anteriorly and '' IRX3'' and ''dbx1a'' posteriorly (genes expressed vary among different organisms), which are

gene In biology, the word gene (from , ; "... Wilhelm Johannsen coined the word gene to describe the Mendelian units of heredity..." meaning ''generation'' or ''birth'' or ''gender'') can have several different meanings. The Mendelian gene is a b ...

s that are expressed in the

and

, respectively. The ZLI is also characterized by a lack of

lunatic fringe Lunatic fringe is a term used to characterize members of a political or social movement as extremists with eccentric or fanatical views. Theodore Roosevelt is widely credited with having popularized the term, which he used with gusto: "There is ap ...

(''lfng''), which is observable even before ''Shh'' expression is observed in the forming ZLI. This indicates that cells are fated for ZLI formation before induction by ''Shh'' (n.b. ''Shh]' signals from the ZLI and is characteristic of the

during and after formation). ''Lfng'' is expressed very early (soon after gastrulation) in the region that will become the ZLI, but shortly thereafter expression recedes through migration of ''lfng''-expressing cells to form the ''lfng''-free wedge characteristic of the developing ZLI. '' Shh'' expression does not extend

until a few hours later.

Positioning

The factors influencing the formation and location of the ZLI are widely studied but still disputed. Differences between different

animal model An animal model (short for animal disease model) is a living, non-human, often genetic-engineered animal used during the research and investigation of human disease, for the purpose of better understanding the disease process without the risk of ha ...

s further complicate elucidation of the

genetic pathway A gene (or genetic) regulatory network (GRN) is a collection of molecular regulators that interact with each other and with other substances in the cell to govern the gene expression levels of mRNA and proteins which, in turn, determine the fun ...

s, specifically between chicks/ mammalian models and

. Wnt (the family wingless) genes are crucial for the development of the ZLI both directly and indirectly in all animal systems. Along with the role of Wnt genes in patterning the

axis through gradient polarization, ''Wnt8b'' is expressed within the ZLI itself and may help guide

dorsal Dorsal (from Latin ''dorsum'' ‘back’) may refer to: * Dorsal (anatomy), an anatomical term of location referring to the back or upper side of an organism or parts of an organism * Dorsal, positioned on top of an aircraft's fuselage * Dorsal c ...

movement of ''Shh'' expression. The Wnt polarization gradient has been linked to induction of ZLI-patterning genes ''IRX3'' and ''

SIX3 Homeobox protein SIX3 is a protein that in humans is encoded by the ''SIX3'' gene. Function The SIX homeobox 3 (SIX3) gene is crucial in embryonic development by providing necessary instructions for the formation of the forebrain and eye deve ...

'', which border the ZLI

posteriorly Standard anatomical terms of location are used to unambiguously describe the anatomy of animals, including humans. The terms, typically derived from Latin or Greek roots, describe something in its standard anatomical position. This position prov ...

and anteriorly, respectively. However, these

s have been shown to be non-essential for ZLI formation in

and have been reevaluated in other models. Specification of the ZLI may also involve the

ventral Standard anatomical terms of location are used to unambiguously describe the anatomy of animals, including humans. The terms, typically derived from Latin or Greek roots, describe something in its standard anatomical position. This position prov ...

tissues of the embryo: the prechordal and epichordal plates, or neuroepithelial tissue (see figure). Interactions between these tissues may be responsible for the recession of expression of ''lfng'' which allows

movement ''Shh'' expression. The specific prechordal and epichordal plates, characterized by expression of ''SIX3'' and ''IRX3'', respectively, may influence positioning of the ZLI more so than the genes themselves. The prechordal plate

ly borders the

, with the epichordal plate posterior to it. Studies of the formation of the ZLI performed in

have revealed the significance of ''

OTX2 Homeobox protein OTX2 is a protein that in humans is encoded by the ''OTX2'' gene. Function This gene encodes a member of the bicoid sub-family of homeodomain-containing transcription factors. The encoded protein acts as a transcription facto ...

'' and ''

IRX1 Iroquois-class homeodomain protein IRX-1, also known as Iroquois homeobox protein 1, is a protein that in humans is encoded by the ''IRX1'' gene. All members of the Iroquois (IRO) family of proteins share two highly conserved features, encoding ...

'' in ZLI positioning. ''OTX2'' expression characterizes the developing

optic tectum In neuroanatomy, the superior colliculus () is a structure lying on the roof of the mammalian midbrain. In non-mammalian vertebrates, the homologous structure is known as the optic tectum, or optic lobe. The adjective form '' tectal'' is commonl ...

, which is responsible for sight processing. Expression extends anteriorly and ends sharply at the ZLI, with high

along the line where ''Shh'' is expressed. Before the ZLI is formed, ''OTX2]'' is expressed ubiquitously throughout the

, and begins to recede to the position of the putative ZLI. Experiments where ''OTX2]'' expression was repressed showed no Dorsum (biology), dorsal movement of ''Shh'' expression and no ZLI formation. ''Irx1'', which is analogous to ''IRX3'' in birds and mammals, is expressed posterior to the ZLI. Studies have suggested that while ''OTX2'' positively restricts the ZLI anteriorly ('' Shh'' cannot be expressed where ''OTX2'' is not), 'IRX1'' negatively restricts the ZLI

. Other genes crucial for differentiation in the brain, including Fgf genes responsible for patterning of the midbrain-hindbrain boundary (MHB), have been implicated in positioning of the ZLI. Studies on the role of ''Shh'' signaling in the ZLI were difficult to study for many years, because mutants lacking expression have many developmental deficits including lack of a diencephalon. Explant and lineage-labeling experiments previously described aided in elucidation of the role of ''Shh'' and other genes in differentiation of these tissues. More recently, the mouse Shh;Gli3 double mutant was found to have an enlarged diencephalon with a ring of Fgf8 and Wnt in place of the ZLI, indicating a complex interaction between Shh and these genes at the ZLI. This also indicates that other patterning cues are able to establish Fgf8 and Wnt signaling domains at the ZLI in the absence of Shh and Gli3.

Differentiation after ZLI degradation

After differentiation of the

progenitor cell A progenitor cell is a biological cell that can differentiate into a specific cell type. Stem cells and progenitor cells have this ability in common. However, stem cells are less specified than progenitor cells. Progenitor cells can only differ ...

s (at a precise

stage Stage or stages may refer to: Acting * Stage (theatre), a space for the performance of theatrical productions * Theatre, a branch of the performing arts, often referred to as "the stage" * ''The Stage'', a weekly British theatre newspaper * Sta ...

yet to be fully determined), the ZLI and its lineage restriction disappears, allowing cells to migrate across the former boundary and the Dorsum (biology), dorsal and

thalami to merge into one functional unit, as shown by replication-incompetent

retroviral A retrovirus is a type of virus that inserts a DNA copy of its RNA genome into the DNA of a host cell that it invades, thus changing the genome of that cell. Once inside the host cell's cytoplasm, the virus uses its own reverse transcriptas ...

experiments that marked cells and showed their migration throughout the diencephalon.

Signaling

After establishment of the ZLI, '' shh'' has been shown to induce

of thalamic and prethalamic markers, ''gbx2'' and ''dlx2''/ ''nkx2.1'', respectively. This differential induction most likely is due to the expression of genes such as ''IRX3'' in the

experiments showed that if ''IRX3'', which is normally expressed in the developing

, is expressed anterior to the ZLI, then the developing

will change identity. Note that these genes which help confer '' shh'' competence help to pattern the ZLI. Signaling from the ZLI cooperating with thalamic and prethalamic markers ensures the migration of post-mitotic (neural progenitor) cells to the mantle zone where the cells assemble into nuclei characteristic of the

. These nuclei are the mechanism of the relay of information from the

to the

cortex Cortex or cortical may refer to: Biology * Cortex (anatomy), the outermost layer of an organ ** Cerebral cortex, the outer layer of the vertebrate cerebrum, part of which is the ''forebrain'' *** Motor cortex, the regions of the cerebral cortex i ...

. The

itself is highly diverse, with each

nucleus Nucleus ( : nuclei) is a Latin word for the seed inside a fruit. It most often refers to: *Atomic nucleus, the very dense central region of an atom * Cell nucleus, a central organelle of a eukaryotic cell, containing most of the cell's DNA Nucl ...

having distinct morphologies and physiologies according to the region of the

to which it is connected. These differences are thought to originate in differential gene expression in the

and

, which allows for a single structure with multiple different and separate functions once the two have merged and completed growth and differentiation. So a functional interplay between Shh from the ZLI and the bHLH factor Her6 (homolog to HES1) determines the neuronal identity within the thalamus: Her6 positive cells in the

and rostral

differentiate into GABAergic inhibitory neurons, whereas her negative cells become

glutamatergic Glutamatergic means "related to glutamate". A glutamatergic agent (or drug) is a chemical that directly modulates the excitatory amino acid (glutamate/aspartate) system in the body or brain. Examples include excitatory amino acid receptor agonis ...

relay neurons. Both cell types depend on Shh signal as trigger to initiate the developmental programme.Scholpp S, Delogu A, Gilthorpe J, Peukert D, Schindler S, Lumsden A. Her6 regulates the neurogenetic gradient and neuronal identity in the thalamus. Proc Natl Acad Sci U S A. 2009 Nov 24;106(47):19895-90

/ref>

References

{{DEFAULTSORT:Zona Limitans Intrathalamica Diencephalon Developmental neuroscience