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One of the greatest challenges in neuroscience research is to understand how biologically active molecules such as small molecules and peptides function at different levels, starting at the atomic level to the ensembles of neuronal networks. Advances made in chemical and synthetic biology over the last few decades have given researchers new insights into bioactive molecular actions. One important key feature that has emerged in recent years is the observation that adult mammalian brains exhibit much more plasticity and regenerative capacity than previously thought. During neurogenesis, the process by which neurons are generated from neural stem and progenitor cells, adult brains are able to generate functionally integrated new neurons. Progenitor cells are ancestor cells that can differentiate into specific cell types. Neuroscientist are just now beginning to study the basic properties of adult neural stem cells and the molecular, cellular and circuitry mechanisms that regulate the sequential adult neurogenesis process in vivo. These studies include the new field called “neuroepigenetics” that investigates the function of novel active DNA modifications and their physiological role in the nervous system.
The discovery of neuropeptides in the mammalian nervous system started a revolution in traditional physiology. Groundbreaking research in physiology, endocrinology, and biochemistry during the last century has let to this discovery. The three notions governing peptide research in the early years of peptide based neuroscience are: (1) peptide hormones are chemical signals in the endocrine system; (2) neurosecretion of peptides is a general principle in the nervous system; and (3) the nervous system is responsive to peptide signals. These ideas have contributed to how neuropeptides are defined today: “Neuropeptides are small substances similar to proteins produced and released by neurons through the regulated secretory route that act on neural substrates.” The beginnings of neuropeptide research can be traced back to the histories of physiology, pharmacology and biochemistry.
Originally the term “neuropeptide” indicated a small protein molecule present in neurons. However, in the late 1970s and the 1980s of the last century, many neuropeptides were localized by immunocyto-chemistry to discrete cell populations of the central and peripheral nervous system leading to the concept of “chemical neuroanatomy” as part of neurobiology. However, recently the field of neuropeptide research or biology has dramatically expanded and today the research goals involve the development of pharmacologically active compounds that are capable of crossing the blood–brain barrier to exert their biological function(s) in vivo and in the construction of genetic vectors useful for gene therapy.
The term ‘neuropeptides’ was first introduced by D. de Wied in 1971 to describe fragments of peptide hormones lacking the activity of the intact hormone molecule but that were able to produce behavioral changes. Subsequently peptide hormones, their fragments, endogenous opioid or morphine-like peptides as well as a large number of other biogenic peptides became classified as neuropeptides. All these peptides are classified based on a number of common features including their origin, biosynthesis, secretion, metabolism, and effectiveness. Neuropeptides are found in the nervous tissue, in peptide-secreting cells present in various organs such as the gut, placenta, heart, lungs, the immune system and other tissues, and are active at extremely low concentrations. The more modern definition of a neuropeptide is as follows: “A neuropeptide is a peptide found in the nervous system which has biological effects when injected.”
Neuropeptides have now been found to be the most diverse class of signaling molecules in the brain that participate in many physiological functions. Almost 70 genes have been identified in the mammalian genome to encode for neuropeptide precursors and a multitude of bioactive neuropeptides. Unfortunately the boundary between a neuropeptide and a neuroprotein can be blurry. This is exemplified in the findings that there are several subfamilies of peptides and small proteins which display most of the hallmarks of neuropeptides. For example among cytokines, peptide hormones, and growth factors there are several subfamilies of peptides that perform similar functions as neuropeptides such as neural chemokines, cerebellins, neurexophilins, and granins. The "cytokine" family alone includes a large and diverse family of regulators produced throughout the body by cells of diverse embryological origin. The use of in situ hybridization and immunocytochemistry allowed scientists to unambiguously demonstrate for many neuropeptides that the transcript and peptide products are produced by neurons. For a few neuropeptides the biosynthesis of the peptides could be demonstrated by the incorporation of radioactive amino acids using pulse-chase experiments.
The recent Annual Neuroscience Meeting held in San Diego from November 9 to 13 demonstrated the importance of all classical neuropeptides as well as of new putative neuropeptides for brain research. The meeting was attended by more than 30,000 neuroscientists and students and the forest of presented posters was so huge that one was only able to visit a selection of them during the meeting hours. One particular peptide that stood out of the crowd is the NAP peptide. This peptide offers new hope to find a cure to prevent the onset of Alzheimers disease or any similar disease, and, hopefully, a few other neuro-degenerative diseases that involve protein unfolding or misfolding events in the mammalian and human brain.
“NAP peptide (Asn-Ala-Pro-Val-Ser-Ile-Pro-Gln) is a neuroprotective peptide that shows cognitive protection in patients with amnestic mild cognitive impairment, a precursor to Alzheimer's disease. NAP exhibits potent neuroprotective properties in several in vivo and cellular models of neural injury. While it has been found in many studies to affect microtubule assembly and/or stability in neuronal and glial cells at fM concentrations, it has remained unclear whether it acts directly or indirectly on tubulin or microtubules. We analyzed the effects of NAP (1 fM-1 µM) on the assembly of reconstituted bovine brain microtubules in vitro and found that it did not significantly (p < 0.05) alter polymerization of either purified tubulin or of a mixture of tubulin and unfractionated microtubule-associated proteins. NAP also had no significant effect (p < 0.05) on the growing and shortening dynamics of steady-state microtubules at their plus ends, nor did it alter the polymerization or dynamics of microtubules assembled in the presence of 3-repeat or 4-repeat tau. Thus, the neuroprotective activity of NAP does not appear to involve a direct action on the polymerization or dynamics of purified tubulin or microtubules.” [Source: Mythili Yenjerla, Nichole E LaPointe, Manu Lopus, Corey Cox, Mary Ann Jordan, Stuart C Feinstein, and Leslie Wilson; The Neuroprotective Peptide NAP Does Not Directly Affect Polymerization or Dynamics of Reconstituted Neural Microtubules. J Alzheimers Dis. 2010 January 1; 19(4): 1377–1386. doi: 10.3233/JAD-2010-1335. PMCID: PMC2844470, NIHMSID: NIHMS179597.]
Neurons use neuropeptides for signaling to other cells. Expression of the genes coding for neuropeptides and their biosynthesis occur in neurons. This has been shown using in situ hybridization and immunocytochemistry. Usually only peptides produced in the nervous system by neurons are called neuropeptides. However, astrocytes and glial cells apparently can also have a regulated secretory pathway and therefore, putative neuropeptides may be recognized in peptide families expressed by glial cells.
Recent research suggest that many neurotransmitters, neuropeptides and calcium binding proteins appear early during development of the cerebellum, have specific temporal and spatial expression patterns, may have functions other than those found in the mature neural systems, and may be able to interact with each other during early development. The basis for regulation of chemical communication is controlled secretion of the signal molecules involved. Neurons expressing classical neuropeptides use the regulated secretory pathway. In this pathway biosynthesized peptides are stored in large dense-cored vesicles and released in a controlled manner upon a stimulus. A signal peptide sequence is needed for entry of the newly synthesized gene product into the lumen of the endoplasmic reticulum (ER) as part of the secretion routes. Usually the signal peptide is short, containing 20 to 25 amino acids at the N-terminal end of the precursor, called the prepro-peptide. This signal peptide is removed from the nascent precursor during protein synthesis when it passes through the ER membrane, leaving the pro-neuropeptide in the ER-Golgi for further sorting into the regulated secretory pathway. Next, neuropeptide precursors are sorted in the trans-Golgi network into the vesicles of the secretory pathway. The resulting protein content is acidified and condensed in the vesicles, and activated proteolytic enzymes process the precursor protein into neuropeptides. Pairs of the basic amino acids lysine (Lys, K) and arginine (Arg, R), or, sometimes, a single Arg in the appropriate structural environment of the precursor, serve as recognition sites and substrates of prohormone convertases (PCs). Next, differential cleavage of the neuropeptide precursor by the proteases leads to the generation of specific peptides and cell-specific different sets of peptides from the same precursor. Peptides generated in this way can be subject to further modification by peptidyl-aminotransferase (PAM). This enzyme uses a C-terminal glycine (Gly, G) as amide donor for the preceding amino acid which results in a peptide with an amidated C-terminus. This amide group has been found to occur in many neuropeptides and is often essential for its biological activity. Furthermore, many neuroppetides have been found to contain post-translational modifications including N- and O-glycosylation, phosphorylation, sulfation, and acetylation. These are found on stored and secreted peptides and contribute to their biological properties. The resulting neuropeptides are stored in mature granules or dense-cored vesicles where they await release upon a stimulus.
Some neuropeptides can also function as hormones. A hormone is defined as a substance secreted by one organ and acting on another after transport by the bloodstream. This allows cells at different locations of an organism to communicate in an endocrine fashion. Since many peptide hormones are also synthesized by neurons these peptides therefore belong to the class of neuropeptides. However, many classical neuropeptides are also synthesized by endocrine glands and function peripherally as hormones which can make the classification of these peptides somewhat confusing. By definition hormones are excluded as being neuropeptides if they are not synthesized by neurons, even if they signal to the brain. Leptin and insulin are examples. In addition, sometimes the relationships between the nervous system and peripheral endocrine glands are complicated by alternative processing and splicing. Hormones are defined as being chemical substances formed or synthesized in one or part of an organ in the body and are carried in the blood to another organ or part. Depending on the specificity or effect of the hormone the targeted organ may be altered in its functional activity or even in its structure by the effect of the hormone. As new peptides are discovered in various tissues that can function as neuropeptides the current classification schemes may need to be expanded.
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